Thermostable Variants of T7 RNA Polymerase

Abstract

A bacteriophage RNA polymerase variant is provided. In some embodiments, the variant may have increased thermostability relative to the corresponding wild type bacteriophage RNA polymerase and/or wild type T7 RNA polymerase. Compositions, kits and methods that employ the variant are also provided.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a continuation of U.S. application Ser. No. 16/680,014, filed on Nov. 11, 2019, which is a continuation of U.S. application Ser. No. 15/594,090, filed on May 12, 2017, which is a continuation-in-part of application No. PCT/US2017/13179, filed on Jan. 12, 2017, which claims the benefit of U.S. provisional application No. 62/416,770, filed Nov. 3, 2016, and U.S. provisional application No. 62/278,161, filed on Jan. 13, 2016, all of which applications are incorporated herein in their entirety by reference.

BACKGROUND

[0002] Thermostable and thermoactive enzymes have great utility in academic research and industrial applications. The high stability of enzymes from thermophilic organisms enables technologies in molecular biology and diagnostics (the Polymerase Chain Reaction, for example). However, equivalent enzymes from thermophilic organisms are not always available. In these cases, directed evolution or computational methods can serve as a powerful tool to identify variants of mesophilic enzymes that confer thermostability. For example, current in vitro transcription methods are limited to reaction temperatures below 45.degree. C. The typical viral RNA polymerases that carry out these reactions are not active at elevated temperatures, and there is a need to identify thermoactive and stable variants in order to carry out in vitro transcription reactions at elevated temperatures.

SUMMARY

[0003] A bacteriophage RNA polymerase variant is provided. In some embodiments, the variant may have increased thermostability and/or activity at elevated reaction temperatures relative to a corresponding wild type RNA polymerase. Compositions, kits and methods that employ the variant are also provided.

[0004] In some embodiments, the variant: (i) comprises an amino acid sequence that has at least 80% (e.g., at least 90%, or at least 95%) sequence identity to SEQ ID NO:1; and (ii) comprises an amino acid substitution at one or more positions corresponding to positions 109, 205, 388, 534, 567 and 618 of SEQ ID NO:1. In some embodiments, the variant may comprise an amino acid substitution at least two positions corresponding to positions 109, 205, 388, 534, 567 and 618 of SEQ ID NO:1. In some embodiments, the variant may comprise an amino acid substitution at least three positions corresponding to positions 109, 205, 388, 534, 567 and 618 of SEQ ID NO:1. In some embodiments, the variant may comprise an amino acid substitution at positions corresponding to positions 109, 205, 388, 534, 567 and 618 of SEQ ID NO:1. For example, in some embodiments, the variant may comprise one or more of the following amino acids substitutions: I109L, H205S, D388E, L534V, V567P and G618Q wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1.

[0005] In one example, variant further includes an amino acid substitution at one or more positions corresponding to positions: 75, 83, 108, 206, 227, 281, 297, 312, 323, 327, 333, 340, 354, 362, 375, 428, 446, 454, 461, 495, 510, 584, 591, 642, 711, 724, 740, 788, 832, 834, 835, 843, 847, 849, 856, 863, 866 and 877 of SEQ ID NO:1.

[0006] In another example, the variant may further comprise an amino acid substitution of at least 10 positions corresponding to positions: 75, 83, 108, 206, 227, 281, 297, 312, 323, 327, 333, 340, 354, 362, 375, 428, 446, 454, 461, 495, 510, 584, 591, 642, 711, 724, 740, 788, 832, 834, 835, 843, 847, 849, 856, 863, 866 and 877 of SEQ ID NO:1.

[0007] In another example, the variant may further comprise one or more of the following amino acids substitutions: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K and E877R, wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1.

[0008] In another example, the isolated bacteriophage RNA polymerase variant, wherein the variant includes at least 10 of the following amino acids substitutions: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K, and E877R, wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1.

[0009] In some embodiments, any isolated bacteriophage RNA polymerase variant described above may include a fusion to an exogenous DNA binding domain. Examples are provided in Table 1. In another embodiment, the variant has increased stability at temperatures of at least 45.degree. C. (e.g., at or above 50.degree. C., or at or above 55.degree. C.) relative the T7 RNA polymerase of SEQ ID NO:1 as a result of the one or more amino acid substitutions.

[0010] Also provided is a composition that includes i. an isolated bacteriophage RNA polymerase variant described above; and ii. a buffering agent. The composition may further include ribonucleoside triphosphates and/or modified nucleotides. The composition may further include a template DNA molecule comprising: a bacteriophage promoter (e.g., a T7 or T3 RNA polymerase promoter) operably linked to a target nucleotide sequence to be transcribed.

[0011] Also provided is a kit is provided that includes i. an isolated bacteriophage RNA polymerase variant of any of those described above; and ii. a reaction buffer. The kit may further comprise one or more ribonucleoside triphosphates and/or modified nucleotides.

[0012] Also provided is a method is provided for synthesizing an RNA molecule that includes

(a) combining an isolated bacteriophage RNA polymerase variant described above with ribonucleoside triphosphates and/or modified nucleotides and a template DNA molecule comprising a bacteriophage RNA promoter that is operably linked to a target nucleotide sequence to be transcribed, to produce a reaction mix; and (b) incubating the reaction mix to transcribe the template DNA molecule into RNA.

[0013] In one embodiment, a method is provided for synthesizing an RNA molecule that includes

(a) incubating the reaction mix described above comprising an isolated bacteriophage RNA polymerase variant described above with ribonucleoside triphosphates and/or modified nucleotides and a template DNA molecule comprising a bacteriophage RNA promoter that is operably linked to a target nucleotide sequence to be transcribed, thereby transcribing the template DNA molecule into RNA.

[0014] In one aspect, the incubating is done at a temperature of at least 45.degree. C. or at above 50.degree. C. or at above 55.degree. C. (for example 45.degree. C. to 60.degree. C., 45.degree. C. to 50.degree. C., 50.degree. C. to 55.degree. C. or 55.degree. C. or 60.degree. C.).

[0015] One example of a bacteriophage RNA polymerase is T7 RNA polymerase.

INCORPORATION BY REFERENCE

[0016] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

[0017] The skilled artisan will understand that the drawings, described below, are for illustration purposes only. The drawings are not intended to limit the scope of the present teachings in any way.

[0018] FIGS. 1A-1D are graphs showing the effect of various amino acid substitutions on the activity of T7 RNA polymerase (SEQ ID NO: 1). FIG. 1A-1C show data for selected variants that have individual mutations. These reactions were done at 45.degree. C. for 10 hours (FIG. 1A and FIG. 1B) and at 37.degree. C. for 2 hours followed by 45.degree. C. for 8 hours (FIG. 1C). FIG. 1D shows the additive effect of individual mutations identified by Tth PURE assay. The reaction was carried out at 45.degree. C. for 10 hours. The thermostability of variants was estimated using the formula, (M-WT)/WT, in which M and WT stand for the maximum value of fluorescent signal from synthesized GFP in 10-hour reactions with mRNA of T7 RNA polymerase variant and wild-type, respectively. In this assay, if a variant polymerase has an activity of "0" then it has the equivalent activity as the wild type T7 RNA polymerase. If a variant polymerase has an activity of "0.5" then it has a 50% increase in activity relative to the wild type T7 RNA polymerase.

[0019] FIGS. 1A-1C show results for 45 single amino acid variants of T7 RNA polymerase.

[0020] FIG. 1D shows the additive effect of combining amino acid substitutions. In FIG. 1D, the additive effect of 1, 2, 3, 4, 5 and 6 amino acids substitutions is shown.

[0021] FIG. 2 shows the melting temperature of wild type T7 RNA polymerase (WT), as well various variants of the same (i.e., M1, M2, M3b, M4, M5, etc.). In this graph, the number after the M corresponds to the number of amino acid substitutions in the polymerase. For example, M5 has five amino acid substitutions relative to wild type T7 RNA polymerase, etc. This data shows that the effect of amino acid substitutions on melting temperature is largely additive.

[0022] FIGS. 3A-3C show that (i) mutant RNA polymerases at temperatures above about 45.degree. C. make more RNA compared to the corresponding wild type; (ii) mutant RNA polymerases at temperatures in the range of 50.degree. C. to 55.degree. C. can make at least 2 fold more RNA than the corresponding wild type RNA polymerase; and (iii) fusion proteins containing RNA polymerase and a DNA binding domain are more active at high temperatures than the corresponding wild type RNA polymerase; and (iv) fusion proteins containing RNA polymerase and a DNA binding domain show prolonged activity with a more gradual loss of activity at temperatures above 56.degree. C. compared to the same RNA polymerase variant alone.

[0023] FIG. 3A shows the transcription activity (RNA synthesis yield) at increased temperature for the wild type T7 RNA polymerase, as well as two variants, commercial T7 RNA polymerase from Toyobo "Toyobo" and M20 where M20 is a mutant of T7 RNA polymerase with 20 amino acid substitutions. As shown, the M20 variant is highly active at temperatures at above 55.degree. C.

[0024] FIG. 3B is a graph showing the transcription activity of wild type T7 polymerase compared to the activity of a variant M18 and a variant M13 and a fusion protein containing the M18 variant and the DNA binding domain of a lacI-like protein from Thermotoga (007) after a 20 minute incubation. As shown the M13 and M18 variants and also the M18 fusion protein is highly active at temperatures at above 55.degree. C. with the fusion protein maintaining its activity (Fluorescence units on the Y axis corresponds to amount of RNA).

[0025] FIG. 3C shows a comparison between wild type, mutant and fusion between mutant and DNA binding domain of a thermostable protein (see Table 1) in which not only is the activity of the fusion variant higher at increasing temperature than the variant alone but also there is slower reduction of activity at temperatures about 56.degree. C. compared to the wild type.

[0026] FIG. 4 is a graph showing the transcription activity of wild type T7 RNA polymerase at different temperatures, compared to the transcription activity of the same T7 RNA polymerase fused to the SS07 DNA binding domain (SS07-T7), the DNA binding domain from a helix-turn-helix (HTH) from Pyrococcus furiosus (109-T7) and the DNA binding domain of a lacI-like protein from Thermotoga (007-T7). The fusion proteins containing T7 RNA polymerase and each of the three DNA binding domain are more active at high temperatures than wild type T7 RNA polymerase.

[0027] FIG. 5 is a graph showing the transcription activity of wild type T7 polymerase compared to the activity of variant M18 which contains 18 amino acid substitutions and a fusion protein containing the M18 variant and the DNA binding domain of a lacI-like protein from Thermotoga (007). Fusion proteins containing thermostable T7 RNA polymerase variants and a DNA binding domain are more active at high temperatures than the thermostable T7 RNA polymerase variants.

[0028] FIG. 6 shows improved thermostability for the T7 RNA polymerase variant identified in SEQ ID NO:70 at 50.degree. C. compared with T7 RNA polymerase wild type under the same reaction conditions.

[0029] FIG. 7A-7B shows that NASBA reactions performed at a higher temperature (e.g., 50-56.degree. C.) have faster amplification times and are more specific than reactions performed at a lower temperature, and that DMSO is not required for this effect. At temperatures below 50.degree. C. all reactions, positive and negative, amplified with similar threshold times. Increasing temperature above 50.degree. C. produced faster amplification times in positive reactions while suppressing amplification in the negative reactions. The no-detectable amplification results are plotted as maximum reaction time, 75 minutes, as indicated by horizontal dashed line. Temperatures of 54-56.degree. C. provided the fastest amplification and maximum discrimination between positive and negative reactions.

[0030] FIG. 7A shows the results of a quantitative NASBA reaction using the M20 variant of T7 RNA polymerase. These reactions do not include DMSO.

[0031] FIG. 7B shows the results of a quantitative NASBA reaction using the M20 variant of T7 RNA polymerase. These reactions include 5% DMSO.

DEFINITIONS

[0032] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Singleton, et al., DICTIONARY OF MICROBIOLOGY AND MOLECULAR BIOLOGY, 2D ED., John Wiley and Sons, New York (1994), and Hale & Markham, THE HARPER COLLINS DICTIONARY OF BIOLOGY, Harper Perennial, N.Y. (1991) provide one of skill with the general meaning of many of the terms used herein. Still, certain terms are defined below for the sake of clarity and ease of reference.

[0033] The term "non-naturally occurring" refers to a composition that does not exist in nature.

[0034] In the context of a protein, the term "non-naturally occurring" refers to a protein that has an amino acid sequence and/or a post-translational modification pattern that is different to the protein in its natural state. For example, a non-naturally occurring protein may have one or more amino acid substitutions, deletions or insertions at the N-terminus, the C-terminus and/or between the N- and C-termini of the protein. A "non-naturally occurring" protein may have an amino acid sequence that is different to a naturally occurring amino acid sequence but that that is at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98% or at least 99% identical to a naturally occurring amino acid sequence. In certain cases, a non-naturally occurring protein may contain an N-terminal methionine or may lack one or more post-translational modifications (e.g., glycosylation, phosphorylation, etc.) if it is produced by a different (e.g., bacterial) cell.

[0035] In the context of a nucleic acid, the term "non-naturally occurring" refers to a nucleic acid that contains: a) a sequence of nucleotides that is different to a nucleic acid in its natural state, b) one or more non-naturally occurring nucleotide monomers (which may result in a non-natural backbone or sugar that is not G, A, T or C) and/or C) may contain one or more other modifications (e.g., an added label or other moiety) to the 5'-end, the 3' end, and/or between the 5'- and 3'-ends of the nucleic acid.

[0036] In the context of a preparation, the term "non-naturally occurring" refers to: a) a combination of components that are not combined by nature, e.g., because they are at different locations, in different cells or different cell compartments; b) a combination of components that have relative concentrations that are not found in nature; c) a combination that lacks something that is usually associated with one of the components in nature; e) a combination that is in a form that not found in nature, e.g., dried, freeze dried, crystalline, aqueous; and/or d) a combination that contains a component that is not found in nature. For example, a preparation may contain a buffering agent (e.g., Tris, HEPES, TAPS, MOPS, tricine or MES), a detergent, a dye, a reaction enhancer or inhibitor, an oxidizing agent, a reducing agent, a solvent or a preservative that is not found in nature.

[0037] As used herein, the term "buffering agent", refers to an agent that allows a solution to resist changes in pH when acid or alkali is added to the solution. Examples of suitable non-naturally occurring buffering agents that may be used in the compositions, kits, and methods of the invention include, for example, Tris, HEPES, TAPS, MOPS, tricine, or MES.

[0038] The term "corresponding to" in the context of corresponding positions, refers to positions that lie across from one another when sequences are aligned, e.g., by the BLAST algorithm.

[0039] The term "variant T7 RNA polymerase" may encompass other types of bacteriophage RNA polymerase with sequences of at least 80% identity to wild type T7 RNA polymerase (SEQ ID NO:1). Enzymes having a similar architecture can be identified using the Conserved Domain Architecture Retrieval Tool (CDART) program of the National Center for Biotechnology Information (Geer, et al. Genome Research 12:1619-1623 (2002)) or by other predictive programs, based on searches employing the sequence of T7 RNA polymerase. Examples of enzymes identified in this manner include: T odd bacteriophages or related viruses including Enterobacteria bacteriophage T7, Yersinia pestis bacteriophage phiA1122; Pseudomonas bacteriophage gh-1; bacteriophage of Pseudomonas putida; Bacteriophage T3; Roseophage S101; and Bacteriophage phiYeO3-12. In addition, other related bacteriophages such as SP6, bacteriophage phiKMV, Enterobacteria bacteriophage K1-5, Vibriophage VpV262, BA14, BA127 and BA156 may encode similar enzymes.

[0040] The term "fusion protein" refers to a DNA binding domain linked to a wild type or variant polymerase. Examples include Pyrococcus furiosus (109-T7) and the DNA binding domain of a lacI-like protein from Thermotoga (007-T7). Other examples are listed in Table 1.

TABLE-US-00001 TABLE 1 DNA binding proteins DNA-binding protein Tfx BD-51 gi|499321160 SEQ ID NO: 10 AbrB/MazE/MraZ-like BD-52 gi|499321199 SEQ ID NO: 11 "Winged helix" DNA-binding domain BD-54 gi|499322061 SEQ ID NO: 12 Ribbon-helix-helix protein, copG family BD-62 gi|499321149 SEQ ID NO: 13 lambda repressor-like DNA-binding domains BD-63 gi|499322443 SEQ ID NO: 14 Resolvase-like BD-67 gi|499322676 SEQ ID NO: 15 "Winged helix" DNA-binding domain BD-71 gi|499322676 SEQ ID NO: 16 "Winged helix" DNA-binding domain BD-74 gi|499322255 SEQ ID NO: 17 "Winged helix" DNA-binding domain BD-75 gi|499322388 SEQ ID NO: 18 "Winged helix" DNA-binding domain BD-81 gi|499322131 SEQ ID NO: 19 "Winged helix" DNA-binding domain BD-82 gi|499321342 SEQ ID NO: 20 "Winged helix" DNA-binding domain BD-85 gi|499321130 SEQ ID NO: 21 "Winged helix" DNA-binding domain BD-86 gi|499322705 SEQ ID NO: 22 "Winged helix" DNA-binding domain BD-88 gi|499320855 SEQ ID NO: 23 "Winged helix" DNA-binding domain BD-89 gi|499322250 SEQ ID NO: 24 "Winged helix" DNA-binding domain BD-91 gi|499321633 SEQ ID NO: 25 "Winged helix" DNA-binding domain BD-92 gi|490170077 SEQ ID NO: 26 "Winged helix" DNA-binding domain BD-93 gi|499321272 SEQ ID NO: 27 "Winged helix" DNA-binding domain BD-94 gi|499320919 SEQ ID NO: 28 "Winged helix" DNA-binding domain BD-97 gi|499320853 SEQ ID NO: 29 "Winged helix" DNA-binding domain BD-98 gi|499321734 SEQ ID NO: 30 "Winged helix" DNA-binding domain BD-100 gi|499322439 SEQ ID NO: 31 "Winged helix" DNA-binding domain BD-102 gi|499322707 SEQ ID NO: 32 "Winged helix" DNA-binding domain BD-109 gi|499321112 SEQ ID NO: 33 HCP-like BD-02 gi|351675391 SEQ ID NO: 34 Helix-turn-helix domain, rpiR family BD-03 gi|500479591 SEQ ID NO: 35 Helix-turn-helix domain, rpiR family BD-04 gi|15643984 SEQ ID NO: 36 Bacterial regulatory proteins, lacI family BD-07 gi|15643711 SEQ ID NO: 37 Bacterial regulatory proteins, lacI family BD-08 gi|15643974 SEQ ID NO: 38 Bacterial regulatory proteins, lacI family BD-09 gi|15643956 SEQ ID NO: 39 Bacterial regulatory proteins, lacI family BD-11 gi|500480095 SEQ ID NO: 40 lambda repressor-like DNA-binding domains BD-12 gi|15643421 SEQ ID NO: 41 "Winged helix" DNA-binding domain BD-14 gi|15644350 SEQ ID NO: 42 "Winged helix" DNA-binding domain BD-16 gi|24159093 SEQ ID NO: 43 "Winged helix" DNA-binding domain BD-18 gi|15643139 SEQ ID NO: 44 "Winged helix" DNA-binding domain BD-23 gi|15642807 SEQ ID NO: 45 "Winged helix" DNA-binding domain BD-24 gi|15643159 SEQ ID NO: 46 "Winged helix" DNA-binding domain BD-30 gi|15643333 SEQ ID NO: 47 "Winged helix" DNA-binding domain BD-32 gi|15643055 SEQ ID NO: 48 "Winged helix" DNA-binding domain BD-37 gi|15643827 SEQ ID NO: 49 "Winged helix" DNA-binding domain BD-43 gi|15643699 SEQ ID NO: 50 Homeodomain-like BD-45 gi|15643788 SEQ ID NO: 51

[0041] The term "temperature-sensitive inhibitor" includes antibody-based hot start RNA polymerase inhibitors where examples of hot start inhibitors for polymerases is provided in Kellogg, et al., Biotechniques, 16(6):1134-7 (1994); aptamer based hot start RNA polymerases where examples for polymerases are provided by New England Biolabs, Ipswich, Mass. (catalog #M0495) and also described by Dang, et al., Journal of Molecular Biology, 264(2), 268-278 (1996); Affibody-based hot start inhibitors where Affibody is a protein-based ligand that inhibits DNA polymerase and exonuclease activity at low temperatures but not at higher temperatures (also described by (Thermo Fisher Scientific, Waltham, Mass. catalog #F549L); and chemical modification resulted in hot start RNA polymerase (see for example U.S. Pat. Nos. 5,773,258; and 6,183,998).

[0042] The term "promoter sequence" includes the sequence 5'-TAATACGACTCACTATAG-3' (SEQ ID NO:74) and any sequence that is at least 90% identical to the canonical sequences for T7. See also Dunn, et al., J Mol Biol. 166(4):477-535 (1983) and Ikeda, et al., J. Biol. Chem. 26, (16): 11322-11328 (1992). This definition also includes the T3 promoter 5' AATTAACCCTCACTAAAG 3' (SEQ ID NO:75) (see New England Biolabs, Ipswich, Mass.) or TATTTACCCTCACTAAAG (SEQ ID NO:76) (Adhya, et al., PNAS 78(1), 147-151 (1981). SP6 promoter has a sequence ATTTAGGTGACACTATAGAAGNG (SEQ ID NO:77) (Thermo Fisher Scientific, Waltham, Mass.). Other promoter sequences are known.

[0043] As used herein, the term "incubating", refers to maintaining a reaction a suitable temperature and time to achieve the desired results, i.e., transcription. Reaction conditions suitable for the enzymes and reagents used in the present method are known (e.g. as described in the Examples herein) and, as such, suitable reaction conditions for the present method can be readily determined. These reactions conditions may change depending on the enzymes used (e.g., depending on their optimum temperatures, etc.).

[0044] As used herein, the term "composition" refers to a combination of reagents that may contain other reagents, e.g., glycerol, salt, dNTPs, etc., in addition to those listed. A composition may be in any form, e.g., aqueous or lyophilized, and may be at any state (e.g., frozen or in liquid form).

DETAILED DESCRIPTION

[0045] Before various embodiments are described in greater detail, it is to be understood that the teachings of this disclosure are not limited to the particular embodiments described, and as such can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present teachings will be limited only by the appended claims.

[0046] While the present teachings are described in conjunction with various embodiments, it is not intended that the present teachings be limited to such embodiments. On the contrary, the present teachings encompass various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art.

[0047] Where a range of values is provided, it is understood that each intervening value, to a half of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the present disclosure.

[0048] Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present teachings, some exemplary methods and materials are now described.

[0049] The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present claims are not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided can be different from the actual publication dates which can need to be independently confirmed.

[0050] It must be noted that as used herein and in the appended claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise. It is further noted that the claims can be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as "solely," "only" and the like in connection with the recitation of claim elements, or use of a "negative" limitation.

[0051] As will be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which can be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present teachings. Any recited method can be carried out in the order of events recited or in any other order which is logically possible.

Variants and Compositions Containing the Same

[0052] Provided herein, in various embodiments, are isolated bacteriophage RNA polymerases belonging to the closely related family of bacteriophage RNA polymerases having at least 80% amino acid sequence identity with T7 RNA polymerase that may be engineered to contain one or more amino acid substitutions corresponding to those identified for T7 RNA polymerase described herein. The isolated bacteriophage RNA polymerase variants may be organized by their improved activity at temperatures of 42.degree. C. and above compared to the corresponding wild type enzyme or wild type T7 RNA polymerase. In some embodiments, the variant: (i) may have an amino acid sequence is at least 80% sequence identity (e.g., at least 90%, at least 90%, at least 95%, at least 97%, at least 98% or at least 99% identity) to SEQ ID NO:1; and (ii) may comprise one or more (e.g., at least two, at least three, at least five, or at least ten) amino acid substitutions at one or more positions corresponding to positions 75, 83, 108, 109, 205, 206, 227, 281, 297, 312, 323, 327, 333, 340, 354, 362, 375, 388, 428, 446, 454, 461, 495, 510, 534, 567, 584, 591, 618, 642, 711, 724, 740, 788, 832, 834, 835, 843, 847, 849, 856, 863, 866, and 877 of SEQ ID NO:1 (wild-type T7 RNA polymerase), shown below:

TABLE-US-00002 SEQ ID NO: 1: MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEAR FRKMFERQLKAGEVADNAAAKPLITTLLPKMIARINDWFEEVKAKRGKRP TAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDEAR FGRIRDLEAKHFKKNVEEQLNKRVGHVYKKAFMQVVEADMLSKGLLGGEA WSSWHKEDSIHVGVRCIEMLIESTGMVSLHRQNAGVVGQDSETIELAPEY AEAIATRAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTH SKKALMRYEDVYMPEVYKAINIAQNTAWKINKKVLAVANVITKWKHCPVE DIPAIEREELPMKPEDIDMNPEALTAWKRAAAAVYRKDKARKSRRISLEF MLEQANKFANHKAIWFPYNMDWRGRVYAVSMFNPQGNDMTKGLLTLAKGK PIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEENHENIMACAKSPLENT WWAEQDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAML RDEVGGRAVNLLPSETVQDIYGIVAKKVNEILQADAINGTDNEVVTVTDE NTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQV LEDTIQPAIDSGKGLMFTQPNQAAGYMAKLIWESVSVTVVAAVEAMNWLK SAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLNLM FLGQFRLQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHE KYGIESFALIHDSFGTIPADAANLFKAVRETMVDTYESCDVLADFYDQFA DQLHESQLDKMPALPAKGNLNLRDILESDFAFA

[0053] For example, in some embodiments, the variant may comprise substitutions at one or more (e.g., one, two, three, four, five or all six) positions corresponding to positions 109, 205, 388, 534, 567 and 618 of SEQ ID NO:1, as well as well as optionally one or more (e.g., at least two, at least three, at least five, or at least ten) other substitutions at other substitutions listed above (see for example FIG. 1A-1D, FIG. 2, FIG. 3A-3C, FIG. 4 and FIG. 5).

[0054] In some embodiments, the isolated T7 RNA polymerase variant: (i) has an amino acid sequence is at least 80% sequence identity (e.g., at least 90%, at least 95%, at least 97%, at least 98% or at least 99% identity) to SEQ ID NO:1; and (ii) comprises one or more (e.g., at least two, at least three, at least five, or at least ten) of the following amino acid substitutions: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K, and E877R, wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1 (see for example FIG. 1A-1D, FIG. 2, FIG. 3A-3C, FIG. 4 and FIG. 5).

[0055] In some embodiments, the variant comprises one or more (e.g., one, two, three, four, five or all six) of the following amino acid substitutions: I109L, H205S, D388E, L534V, V567P and G618Q, wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1, as well as well as optionally one or more (e.g., at least two, at least three, at least five, or at least ten) of the following amino acid substitutions: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K, and E877R, wherein the amino acid substitutions are at positions that correspond to positions in SEQ ID NO:1 (see for example FIG. 1A-1D, FIG. 2, FIG. 3A-3C, FIG. 4 and FIG. 5).

[0056] As would be apparent, RNA polymerase variants described herein have RNA polymerase activity and, as such, can catalyze the formation of RNA in the 5'.fwdarw.3' direction using a DNA template. T7 RNA polymerase is a promoter-specific polymerase that transcribes downstream of a suitable promoter (e.g., TAATACGACTCACTATAG; SEQ ID NO:2). In certain embodiments, the non-natural bacteriophage RNA polymerase may activate transcription from a promoter that has at least 90% sequence identity with SEQ ID NO:2; AATTAACCCTCACTAAAG (SEQ ID NO:3); TATTTACCCTCACTAAAG (SEQ ID NO:4) or ATTTAGGTGACACTATAGAAGNG (SEQ ID NO:5). Transcription typically beginnings at the 3' G nucleotide. The polymerase variants also preferably utilize Mg.sup.2+ ion as cofactor for the synthesis of RNA.

[0057] T7 RNA polymerase which is generally described in Maslak et al, Biochemistry 1994, 33: 6918-6924; Martin et al Prog. Nucleic Acid Res. Mol. Biol. 2005 80: 323-47; and Sousa et al Prog. Nucleic Acid Res. Mol. Biol. 2003 73: 1-41, is structurally related to other bacteriophage polymerases such as T3 polymerase (NP_523301.1) and SP6 polymerase (YP_004306655.1) as well as the RNA polymerases from Yersinia bacteriophage phiYeO3-12, Erwinia bacteriophage FE44, Kluyvera bacteriophage Kvp1, Enterobacteria bacteriophage K1F, Vibrio bacteriophage ICP3_2009_A and Pseudomonas bacteriophage PPPL-1. It is expected that the amino add substitutions described herein may be transferred to other, related RNA polymerases and their variants with the same effect. As such, in certain embodiments, this disclosure provides a non-naturally occurring variant of a naturally occurring bacteriophage RNA polymerase, wherein the variant has an amino acid that is at least 80% identical to (e.g., at least 90%, at least 95% or at least 98% identical to) the naturally occurring bacteriophage RNA polymerase and comprises one or more amino acid substitutions relative the naturally occurring to bacteriophage RNA polymerase, wherein the one or more amino acid substitutions are at one or more position listed above.

[0058] In some embodiments, an isolated bacteriophage RNA polymerase variant with one or more amino acid substitutions has increased stability at 45.degree. C. or greater relative to the wild type RNA polymerase. This is here exemplified by a T7 RNA polymerase variant, having one or more amino acid substitutions, and by a fusion between the bacteriophage RNA polymerase and a DNA binding protein such as described in Table 1 and in Example 3. In some embodiments, an isolated bacteriophage RNA polymerase variant exemplified by a T7 RNA polymerase variant may be used in an in vitro transcription reaction that is incubated at an elevated temperature (e.g., a temperature in the range of for example, 45.degree. C. to 60.degree. C., 45.degree. C. to 50.degree. C., 50.degree. C. to 55.degree. C. or 55.degree. C. or 60.degree. C.) to produce at least 10% more product, at least 20% more product, at least 50% more product, at least 100% more product, or at least 500% more product than an otherwise identical reaction containing the wild type RNA polymerase (e.g. SEQ ID NO:1 for T7 RNA polymerase) incubated under the same conditions.

[0059] Also provided is a composition, e.g., an aqueous composition comprising: i. an isolated bacteriophage RNA polymerase variant (e.g., a T7 RNA polymerase described herein) and ii. a buffering agent (e.g., Tris). In some embodiments, the composition may be a composition in which the polymerase can be stored. In these embodiments, the composition may optionally contain glycerol, salt (e.g., NaCl), EDTA, detergent (e.g., Triton X-100). In other embodiments, the composition may be a reaction mix. In these embodiments, the composition may further comprise ribonucleoside triphosphates (e.g., one, two, three or all four of ATP, UTP, GTP, CTP) and/or one or more modified nucleotides. In some embodiments, the composition may further comprise a template DNA molecule comprising: a bacteriophage promoter operably linked to a target nucleotide sequence to be transcribed. In some embodiments, the composition may comprise a population of such template DNA molecules, where each of the template molecules comprises a bacteriophage promoter upstream from a target nucleotide sequence to be transcribed. The bacteriophage promoter can be any of those described herein such as a T7 promoter, a T3 promoter or an SP6 promoter. A reaction mix composition may additionally contain 4-10 mM MgCl.sub.2, e.g., 6 mM, MgCl.sub.2.

[0060] In some embodiments, a variant may be fused to a DNA binding domain, the activity of the RNA polymerase may be enhanced at elevated temperatures by 50% or 100% or 150% or 200% or more (see for example, FIG. 4 or FIG. 5).

[0061] Kits

[0062] Also provided is a kit comprising: i. an isolated bacteriophage RNA polymerase variant as described herein; and ii. a reaction buffer. In some embodiments, the kit may further comprise one or more ribonucleoside triphosphates (e.g., one, two, three or all four of ATP, UTP, GTP, CTP). The components of the kit may be combined in one container, or each component may be in its own container. For example, the components of the kit may be combined in a single reaction tube or in one or more different reaction tubes. Further details of the components of this kit are described above. The kit may also contain other reagents described above and below that may be employed in the method depending on how the method is going to be implemented. In some embodiments, the kit may comprise of a variant as described above and a buffer in which the variant is active, or a concentrated form thereof.

[0063] In addition to above-mentioned components, the subject kit may further include instructions for using the components of the kit to practice the subject method. The instructions for practicing the subject method are generally recorded on a suitable recording medium. For example, the instructions may be printed on a substrate, such as paper or plastic, etc. As such, the instructions may be present in the kits as a package insert, in the labeling of the container of the kit or components thereof (i.e., associated with the packaging or subpackaging) etc. In other embodiments, the instructions are present as an electronic storage data file present on a suitable computer readable storage medium, e.g. CD-ROM, diskette, etc. In yet other embodiments, the actual instructions are not present in the kit, but means for obtaining the instructions from a remote source, e.g. via the internet, are provided. An example of this embodiment is a kit that includes a web address where the instructions can be viewed and/or from which the instructions can be downloaded. As with the instructions, this means for obtaining the instructions is recorded on a suitable substrate.

[0064] Methods

[0065] Also provided is a method for synthesizing an RNA molecule. In some embodiments, this method may comprise (a) combining an isolated bacteriophage RNA polymerase variant described herein with ribonucleoside triphosphates and/or modified nucleotides and a template DNA molecule comprising a promoter operably linked to a target nucleotide sequence to be transcribed, to produce a reaction mix; and (b) incubating the reaction mix to transcribe the template DNA molecule into RNA. In some embodiments, the incubating may be done at a temperature of at least 45.degree. C. (e.g., in the range of 45.degree. C. to 60.degree. C., 45.degree. C. to 50.degree. C., 50.degree. C. to 55.degree. C. or 55.degree. C. or 60.degree. C.) to transcribe the DNA into RNA. The DNA can be single- or double-stranded and should have a promoter recognized by the polymerase. In one embodiment, the method includes a T7 RNA polymerase variant and a T7 promoter or T3 promoter or variants thereof.

[0066] In some embodiments, the present RNA polymerase may be used to amplify RNA by NASBA (Nucleic Acid Sequence Based Amplification). NASBA was initially described by Compton (Nature, 350 (6313):91-92 (1991)) and has been used as a rapid diagnostic tests for several pathogenic viruses with RNA genomes, e.g. influenza A, foot-and-mouth disease virus, severe acute respiratory syndrome (SARS)-associated coronavirus, human bocavirus (HBoV) and also parasites like Trypanosoma brucei as well as other viruses such as HIV-1 (see, e.g., Kievits Journal of Virological Methods. 1991 35: 273-86). NASBA can be used for medical diagnostics, where it has been shown to give quicker results than PCR, and it can also be more sensitive. NASBA's is an isothermal reaction that is typically run at a constant temperature of at least 41.degree. C. When a present variant is used, the incubation temperature can be increased to above at least 45.degree. C. (e.g., in the range of 45.degree. C. to 60.degree. C., 45.degree. C. to 50.degree. C., 50.degree. C. to 55.degree. C. or 55.degree. C. or 60.degree. C.). In some implementations, when the RNA template is added the reaction mixture, a primer containing a promoter sequence hybridizes to a complementary site at the 3' end of the template, and reverse transcriptase synthesizes the opposite, complementary DNA strand. RNAse H destroys the RNA template from the DNA-RNA hybrid, and a second primer hybridizes to the 5' end of the cDNA strand. The second primer is extended using the cDNA as a template, resulting in double stranded DNA. A T7 RNA polymerase variant can continuously produce complementary RNA strands of this template, which results in amplification. The amplicons, however, are antisense to the original RNA template. A higher incubation temperature results in less non-specific binding of DNA primers to the RNA. In some embodiments, the reaction may contain temperature-sensitive inhibitor of the polymerase, thereby allowing the polymerase to remain inactive until the temperature is raised. In other embodiments, the present RNA polymerase may also be used to amplify RNA by TMA (Transcription-Mediated Amplification). TMA is an isothermal, single-tube nucleic acid amplification system utilizing two enzymes, RNA polymerase and reverse transcriptase, to rapidly amplify the target RNA/DNA, enabling the simultaneous detection of multiple pathogenic organisms in a single tube. TMA technology allows a clinical laboratory to perform nucleic acid test (NAT) assays for blood screening with fewer steps, less processing time, and faster results. It is used in molecular biology, forensics, and medicine for the rapid identification and diagnosis of pathogenic organisms. In contrast to similar techniques such as polymerase chain reaction and ligase chain reaction, this method involves RNA transcription (via RNA polymerase) and DNA synthesis (via reverse transcriptase) to produce an RNA amplicon (the source or product of amplification) from a target nucleic acid. This technique can be used for both target RNA and DNA. NASBA and TMA reactions that are performed at a higher temperature (e.g., in the range of 45.degree. C. to 60.degree. C.) produce less non-specific amplification products relative to the same reactions that are done at a lower temperature (e.g., 41.degree. C.). Moreover, DMSO or other compounds that lower the melting temperature of the primers and substrate DNA or RNA do not need to be added to the amplification reaction at a higher temperature using a thermostable RNA polymerase. As such, in some embodiments, amplification reactions (e.g., a NASBA or TMA reactions) that are done using a thermostable RNA polymerase may be done using a reaction mix that does not contain DMSO or a functional equivalent thereof (e.g, betaine, ethylene glycol and 1,2-propanediol, etc.)

[0067] In some embodiments, the RNA product (the product of an amplification reaction may be detected after it is amplified. This may be done, e.g., using a molecular beacon (see, e.g., Tyagi Nat. Biotechnol. 1996 14: 303-8, among others). Detection may be done quantitatively in some cases.

[0068] Examples of closely related bacteriophage RNA polymerases are provided below. Mutations identified for T7 RNA polymerase that improve thermostability and/or activity are expected to have a corresponding effect when positioned in closely related bacteriophage RNA polymerases in corresponding positions.

TABLE-US-00003 Enterobacteria bacteriophage 13a (SEQ ID NO: 52) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKMFERQLKAGEVADNAAAKPLI- TTLLPKMIARIN DWFEEVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSVDNTTVQAVASAIGRAIEDEARFGRIRDLEAK- HFKKNVEEQLNKR VGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIESTGMVSLHRQNAGVVGQDSETIELA- PEYAEAIAT RAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWKI- NKKVLAVA NVITKWKHCPVEDIPAIEREELPMKPEDIDTNPDALTAWKRAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- AIWFPYNMD WRGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKS- PLENTWWA EQDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLLDEIGGRAVNLLPSETVQDIYGIVAKK- VNVILQADVINGT DNEVVTVTDENTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLN- LMFLGQFR LQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAANLFKAVRE- TMVDTYESCDVL ADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Yersinia bacteriophage YpP-R (SEQ ID NO: 53) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKMFERQLKAGEVADNAAAKPLI- TTLLPKMIARIN DWFEEVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDEARFGRIRDLEAK- HFKKNVEEQLNKR VGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIESTGMVNLHRQNAGVVGQDSETIELT- PEYAEAIAT RAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWKI- NKKVLAVA NVITKWKHCPVEDIPAIEREELPMKPEDIDTNPEALTAWKRAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- AIWFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKSP- LENTWWAE QDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLLDEVGGLAVNLLPSATVQDIYGIVAKKV- NVILQADVINGT DNEVVTVTDENTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLN- LMFLGQFR LQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAANLFKAVRE- TMVDTYESCDVL ADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Yersinia bacteriophage R (SEQ ID NO: 54) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKMFERQLKAGEVADNAAAKPLI- TTLLPKMIARIN DWFEEVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDEARFGRIRDLEAK- HFKKNVEEQLNKR VGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIESTGMVNLHRQNAGVVGQDSETIELT- PEYAEAIAT RAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWKI- NKKVLAVA NVITKWKHCPVEDIPAIEREELPMKPEDIDTNPEALTAWKRAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- AIWFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKSP- LENTWWAE QDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLLDEVGGLAVNLLPSATVQDIYGIVAKKV- NVILQADVINGT DNEVVTVTDENTGEIPEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLN- LMFLGQFR LQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAANLFKAVRE- TMVDTYESCDVL ADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Yersinia bacteriophage phiA1122 (SEQ ID NO: 55) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKMFERQLKAGEVADNAAAKPLI- TTLLPKMIARIN DWFEEVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDEARFGRIRDLEAK- HFKKNVEEQLNKR VGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIESTGMVNLHRQNAGVVGQDSETIELT- PEYAEAIAT RAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWKI- NKKVLAVA NVITKWKHCPVEDIPAIEREELPMKPEDIDTNPEALTAWKRAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- AIWFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKSP- LENTWWAE QDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLLDEVGGLAVNLLPSATVQDIYGIVAKKV- NVILQADVINGT DNEVVTVTDENTGEIPEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLN- LMFLGQFR LQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAANLFKAVRE- TMVDTYESCDVL ADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Escherichia bacteriophage CICC 80001 (SEQ ID NO: 56) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKMFERQLKAGEVADNAAAKPLI- TTLLPKMIARIN DWFEVVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDESRFGRIRDLEAK- HFKKNVEEQLNKR VGHVYRKAFMQVVEADMLSKGLMGGEAWSSWHKEDSIHVGVRCIEMLIESTGMVSLHRQNAGVVGQDSETIELA- PEYAEAIA TRAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWK- INKKVLAVA NVITKWKHCPVEDIPAIEREELPMKPEDIDTNPEALTAWKRAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- SIWFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKSP- LENTWWAE QDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGLAVNLLPSETVQDIYGIVAKKV- NVILQEDVINGT DNEVVTVTDENTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLN- LIFLGQFRL QPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAANLFKAVRET- MVDTYESCDVLA DFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Yersinia bacteriophage YpsP-G (SEQ ID NO: 57) MTERTDGLKKGYMPNGTLYAANRRLVRTWRENNLELAAIPFNTLADHYGERLAREQLALEHESYEMGEARFRKM- FERQLKAGE VADNAAAKPLITTLLPKMIARINDWFEEVKAKRGKRPTAFQFLQEIKPEAVAYITIKTTLACLTSADNTTVQAV- ASAIGRAIEDEARF GRIRDLEAKHFKKNVEEQLNKRVGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIESTG- MVNLHRQN AGVVGQDSETIELTPEYAEAIATRAGALAGISPMFQPCVVPPKPWTGITGGGYWANGRRPLALVRTHSKKALMR- YEDVYMPEVY KAINIAQNTAWKINKKVLAVANVITKWKHCPVEDIPAIEREELPMKPEDIDTNPEALTAWKRAAAAVYRKDKAR- KSRRISLEFMLE QANKFANHKAIWFPYNMDWRGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYYWLKIHGANCAGVDKVPFPE- RIKFIEDN HENIMACAKSPLENTWWAEQDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLLDEVGGLAV- NLLPSATVQD IYGIVAKKVNVILQADVINGTDNEVVTVTDENTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVMTLAYGSKE- FGFRQQVLEDTI QPVIDSGKGLMFTQPNQAAGYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTGElLRKRCAVHWVT- PDGFPVW QEYKKPIQTRLNLMFLGQFRLQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALI- HDSFGTIPADAA NLFKAVRETMVDTYESCDVLADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Salmonella bacteriophage Vi06 (SEQ ID NO: 58) MNTISITKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEVRFRKMFERQLKAGEIADNDATKPLI- TTLLPKMIARINS WFKEVQAKCGKRPTAFQFLQGIKPEAIAYITIKTTLARLTSMDNTTVQAVASAIGRAIEDEARFGRIRDLEAKH- FKKNVEEQLNKRV GHVYKKAFMQVIEADMLSKGLLGGESWSSWHKEDSIHVGVRCIEMLIESTGMVSLHRQNAGVVGQDSETIELAP- EYAEAIATRA GALAGISPMFQPCVVPPKPWTSISGGGYWANGRRPLALVRTHSKKALMRYADVYMPEVYKAVNIAQNTAWRINK- KVLAVANV VTKWKHCPVDYIPTIEREELPMKPEDIDTNPEALASWKRAAAAVYRKDKARKSRRMSLEFMLEQANKFANHRAI- WFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGFYWLKIHGANCAGVDKVPFPERIKFIEDNHENILACAKSP- LENTWWSEQ DSPFCFLAFCFEYAGGQHHGLSYNCSLPLAFDGSCFGIQHFSVMLRDEVGGRAVNLLPSETVQDIYGIVAKKVN- EILQVDMINGT DNEVVTVTDDKTGEIYEKIKLGTKELAGQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTHPNQAA GYMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAVEVKDRKTGElLRKRCAVHWTTPDGFPVWQEYKKPVQTRLN- LIFLGQFRL QPTINTNRDSEIDAYKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIDSFALIHDSFGTIPADAANLFKAVRET- MVATYESCDVLA DFYAQFADQLHKSQLDKMPVLPSKGNLNLQDILKSDFAFA Stenotrophomonas bacteriophage IME15 (SEQ ID NO: 59) MTVIAIEKNDFSDVELAVIPFNTLADHYGEKLAREQLALEHEAYEMGEARFRKIFERQLKAGEVADNAAAKPLV- ATLLPKMIERIHA WFEEVSAKRGKRPTAFKFLQEVKPEAIAYITIKTVLGTLTSAEQTTVQAAASAVGRAIEDEARFGRIRDLEAKH- FKKNVEEQLNKRV GHVYKKAFLQVVEADMLSKGLMGGEAWSSWHKEDSIHVGVRCIEMLIEATGLVVLERQNAGVVGADAETLSLAS- EYADAIATR AGALAGISPMYQPCVVPPKPWTTVTGGGYWANGRRPLALVRTHGKKALMRYEDVYMPEVYKAVNLAQSTAWKIN-

KKVLAVA NEITKWKHCPVEDIPAIEREELPVKPDDIDENPEALTNWKRAAAAVYRKDKARKSRRLSLEFMLEQANKFANHK- AIWFPYNMDW RGRVYAVSMFNPQGNDMTKGLLTLAKGKAIGKEGFYWLKIHGANCAGVDKVPFPERIKFIEDNHEHIMASAKNP- LEYTWWAE QDSPFCFLAFCFEYAGVMHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAKKV- NEIMQRDVISG TDDELVTETDKTTGEITEKAVLGTRTLAGQWLAYGANRSVTKRSVMTLAYGSKEFGFRQQVLEDTIRPAIDSGK- GLMFTIPNQAA GYMAKLIWDSVSVTVVAAVEAMKWLQSAAKLLAAEVKDKKTGEVLRNRCAVHWVTPDGFPVWQEYRKPLQTRLN- LMFLGQF RLQPTINTNKDSGIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHDSFGTIPADAGNLFKAVR- ETMVDTYENCD VLADFYEQFADQLHESQLDKMPALPKKGNLNLRDILESDFAFA Citrobacter bacteriophage SH2 (SEQ ID NO: 60) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYASKKGRKPVAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVSITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGVYRLDKARVSRRISLEFMLEQANKFANKK- AIWFPYNMD WRGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKD- PINNTWWAE QDSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKV- NEILKQDAINGT PNEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLD- MIFLGQFR LQPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRE- TMVITYENNDVL ADFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Enterobacter bacteriophage E-4 (SEQ ID NO: 61) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYASKKGRKPSAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVSITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFANKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- IFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRET- MVLTYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Yersinia bacteriophage phiYe-F10 (SEQ ID NO: 62) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYASKKGRKPVAYAPLQSLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVSITGGGYWANGRRPLALIRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGVYRLDKARVSRRISLEFMLEQANKFASKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- IFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRET- MVITYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Citrobacter bacteriophage phiCFP-1 (SEQ ID NO: 63) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYDSKKGRKPVAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVAITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFASKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- IFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRET- MVITYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Citrobacter bacteriophage SH1 (SEQ ID NO: 64) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYASKKGRKPVAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVSITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFASKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLADEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- lFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRET- MVITYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Salmonella bacteriophage phiSG-JL2 (SEQ ID NO: 65) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTARIVE WLEEYASKKGRKPVAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVAITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGVYRLDKARVSRRISLEFMLEQANKFASKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- IFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRET- MVLTYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGKLNLQDILKSDFAFA Yersinia bacteriophage phiYeO3-12 (SEQ ID NO: 66) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAREQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTTRIVE WLEEYATKKGRKPVAYAPLQSLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVAITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFASKK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKDP- INNTWWAEQ DSPFCFLAFCFEYAGVAHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKVN- ElLKQDAINGTP NEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKGL- MFTQPNQAAG

YMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLDM- IFLGQFRL QPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHENYGIESFALIHDSFGTIPADAGKLFKAVRET- MVITYENNDVLA DFYDQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Enterobacteria bacteriophage T7M (SEQ ID NO: 67) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAKEQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTTRIVE WLEEYASKKGRKPSAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVAITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNLAQNTAWKI- NKKVLAV VNEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFASK- KAIWFPYNMD WRGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKD- PINNTWWAE QDSPFCFLAFCFEYAGVTHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKV- NEILKQDAINGT PNEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLD- MIFLGQFR LQPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRE- TMVITYENNDVL ADFYSQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Enterobacteria bacteriophage T3 (SEQ ID NO: 68) MNIIENIEKNDFSEIELAAIPFNTLADHYGSALAKEQLALEHESYELGERRFLKMLERQAKAGEIADNAAAKPL- LATLLPKLTTRIVE WLEEYASKKGRKPSAYAPLQLLKPEASAFITLKVILASLTSTNMTTIQAAAGMLGKAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRH GQVYKKAFMQVVEADMIGRGLLGGEAWSSWDKETTMHVGIRLIEMLIESTGLVELQRHNAGNAGSDHEALQLAQ- EYVDVLAK RAGALAGISPMFQPCVVPPKPWVAITGGGYWANGRRPLALVRTHSKKGLMRYEDVYMPEVYKAVNLAQNTAWKI- NKKVLAV VNEIVNWKNCPVADIPSLERQELPPKPDDIDTNEAALKEWKKAAAGIYRLDKARVSRRISLEFMLEQANKFASK- KAIWFPYNMD WRGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGEEGFYWLKIHGANCAGVDKVPFPERIAFIEKHVDDILACAKD- PINNTWWAE QDSPFCFLAFCFEYAGVTHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAQKV- NEILKQDAINGT PNEMITVTDKDTGEISEKLKLGTSTLAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLDDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRHRCAVHWTTPDGFPVWQEYRKPLQKRLD- MIFLGQFR LQPTINTLKDSGIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGKLFKAVRE- TMVITYENNDVL ADFYSQFADQLHETQLDKMPPLPKKGNLNLQDILKSDFAFA Phage RNA polymerase (SEQ ID NO: 69) MNIINIAKNDFSDIELAAIPFNILADHYGAQLAREQLALEHEAYEEGEKRFLKMLERQIKAGEFADNAAAKPLL- STLLPKLIARINDW FEEVAAKRGKKPVAYNPLQHVKPEAAAFITLKVTLACLTKAEFTTIQAVASAIGRAIEDEARFGRIRDLEAKHF- KKHVEEQLNKRVG HVYKKAFMQVVEADMLSKGLLGGEAWSSWTKEESIHVGVRMLELLIESTGLVELHRPNAGNVGKDVEMIQLAPE- YVDLLAKRA GALAGISPMYQPCVVPPKPWTSIVGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAVNIAQNTPWKINK- KVLAVVNEI VNWKHCPVADVPAIEREELPPKPEDIDTNEAALKAWKKAAAAIYRKDKARVSRRLSMEFMLEQANKFANFKAIW- FPYNMDWR GRVYAVPMFNPQGNDMTKGLLTLAKGKPIGKDGFYWLKIHGANCAGVDKVPFPERIKFIEDNHENIMACAKDPL- NNEWWAEQ DSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEIGGRAVNLLPSETVQDIYGIVADKVN- EILKQDAINGTD NEVETVTDKDTGEITEKLKLGTKELAGQWLAYGVTRKVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKGL- MFTQPNQAAG YMAKLIWEAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEVLRKRCAVHWVTPDGFPVWQEYRKPVQTRLNL- MFLGQFRL QPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRMTVVHAHEKYGIESFALIHDSFGTIPADAGNLFKAVRET- MVNTYEDNDVL ADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA Phage RNA polymerase (SEQ ID No: 70) MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYEMGEKRFLKMLERQVKAGEIADNAAAKPLI- TTLLPKLTARIND WFEEVAAKRGKRPVAYQPLQGIKPEAVAFITIKVVLASLTSADNTTIQAVASAIGRAIEDEARFGRIRDLEAKH- FKKHVEEQLNKRV GHVYKKAFMQVVEADMLSKGLLGGEAWSSWNKEESMHVGIRMIEMLIESTGLVELHRHNAGVVGQDSETIQLAP- EYVEALAK RAGALAGISPMFQPCVVPPKPWVSITGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAVNIAQNTAWKI- NKKVLAVV NEIVNWKHCPVEDIPAIEREELPPKPDDIDTNEEALKAWKKAAAAVYRKDKARKSRRISLEFMLEQANKFANHK- AIWFPYNMDW RGRVYAVPMFNPQGNDMTKGLLTLAKGKPIGKEGFYWLKIHGANCAGVDKVPFPERIKFIEDNHDNIMACAKDP- LDNTWWAE QDSPFCFLAFCFEYAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVADKV- NEILKQDVINGT DNEVVTVTDKDTGEISEKLKLGTKELAQQWLAYGVTRSVTKRSVMTLAYGSKEFGFRQQVLEDTIQPAIDSGKG- LMFTQPNQAA GYMAKLIWDAVSVTVVAAVEAMNWLKSAAKLLAAEVKDKKTKEILRKRCAVHWVTPDGFPVWQEYRKPIQTRLN- LMFLGQFR LQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRMTVVYAHEKYGIESFALIHDSFGTIPADAGNLFKAVRE- TMVNTYENNDV LADFYDQFADQLHESQLDKMPALPAKGNLNLQDILKSDFAFA

EXAMPLES

Example 1: Initial Screening Assays

[0069] Structure-based calculations were carried out to predict effect of mutations on thermal stability of T7 RNA polymerase. Mutations were modeled and evaluated using BioLuminate.TM. software (Schrodinger, New York, N.Y.) starting with the crystal structure of the wild-type T7 RNA polymerase (PDB ID: 1MSW). The predicted change in protein thermal stability upon mutation (MG) was used to choose the candidate mutations. Mutations predicted to be stabilizing were introduced into wild type T7 RNA polymerase using site-directed mutagenesis using the Q5.RTM. Site-Directed Mutagenesis Kit (New England Biolabs, Ipswich, Mass.) and manufacturer's recommended protocols.

[0070] Individual mutations (see for example those in FIGS. 1A-1D) were screened in a novel cell-free assay based on the reconstituted translation system from Thermus thermophilus (Tth PURE system). Reconstitution of translation from Thermus thermophilus reveals a minimal set of components sufficient for protein synthesis at high temperatures and functional conservation of modern and ancient translation components (Zhou, et al., Nucleic Acids Research, 40(16), 7932-7945 (2012)). Genes encoding T7 RNAP variants were transcribed in vitro using SP6 RNA polymerase. 1 .mu.l of in vitro transcription reaction was added to 10 .mu.l of Tth PURE system with a fluorescent reporter gene (a GFP variant under the control of a T7 RNAP promoter). The activity of T7 RNA polymerase variants synthesized in Tth PURE system was coupled to the expression of a GFP gene under the control of a T7 promoter. All reactions were incubated at a range of temperatures for 10 hours. Transcription was monitored by production of a fluorescent signal in real time using a CFX96 Touch.TM. Real-Time PCR Detection System (Bio-Rad, Hercules, Calif.). FIG. 1A-1C show data for selected individual variants incubated at 45.degree. C. for 10 hours (FIGS. 1A and 1B) and at 37.degree. C. for 2 hours followed by 45.degree. C. for 8 hours (FIG. 1C). FIG. 1D shows data for selected combinations of mutations. The reaction was carried out at 45.degree. C. for 10 hours. All variants shown have a detectable increase in thermostability.

Example 2: Melting Temperature and Temperature Dependence Analysis

[0071] A. Selected T7 RNA polymerase variants (including those described in FIGS. 3A-3C, and FIGS. 4-6) contained an N-terminal hexahistidine tag and were expressed in E. coli and purified using nickel affinity chromatography using an AKTAFPLC.RTM. system (GE Life Sciences, Marlborough, Mass.). The hexahistidine-tagged polymerase variants were isolated and purified on nickel resin, eluted from the nickel resin with imidazole and dialyzed into a storage buffer (for example: 50 mM Tris-HCl pH 7.5, 100 mM NaCl, 10 mM DTT, 50% Glycerol, 0.1% Triton X-100).

[0072] B. To measure the melting temperature of the mutants (as shown in FIG. 2), 0.2 mg/ml T7 RNA polymerase variants were prepared in a buffer (50 mM Hepes-KOH, pH 8.0, 10 mM Mg(OAc).sub.2, 5 mM DTT, 2 mM spermidine). Melting temperatures were measured using Prometheus NT.48 (NanoTemper Technologies).

[0073] C. To determine the reaction temperature range (as shown in FIG. 3A), the yield of RNA synthesis was measured from 37.degree. C. to 60.degree. C. Each 25 .mu.l reaction contains a final concentration of 50 mM Hepes-KOH, pH 7.5, 10 mM Mg(OAc).sub.2, 5 mM DTT, 2 mM spermidine, 1 mM NTP, 4 ng/.mu.l linear DNA template of the Green Fluorescent protein reporter gene with T7 promoter, and 8 .mu.g/ml T7 RNA polymerase variant. Reactions were run at various temperatures using Bio-Rad T100.TM. Thermal Cycler (Bio-Rad, Hercules, Calif.) for 2 hours. After the transcription reactions, 1 unit of DNase I (New England Biolabs, Ipswich, Mass.) was added and the reactions were incubated at 37.degree. C. for 30 minutes.

[0074] Total synthesized RNA was measured using a Qubit.RTM. RNA BR Assay Kit (Thermo Fisher Scientific, Waltham, Mass.) to measure GFP mRNA.

Example 3: Beacon Assays

[0075] Wild type T7 RNA polymerase and two different variants were fused to the sso7d DNA binding domain (of sequence ATVKFKYKGEEKEVDISKIKKVWRVGKMISFTYDEGGDKTGRGAVSEKDAPKELLQMLEKQKK; SEQ ID NO:6), the DNA binding domain from a helix-turn-helix (HTH) from Pyrococcus furiosus (of sequence GRKVRTQQNEILNLLNEKEKAVLRAILEHGGEIKQED LPELVGYSRPTISKVIQELENKGLIKREKSGKTFVVKIERKIKLDKMGAPT; SEQ ID NO:7) or the DNA binding domain of a lacI-like protein from Thermotoga (of sequence KRRPTINDVAKLAGVSISTVSRYLKDPSQVSEKLGERIREAIKKLGYKPNKIAQGLRTGD; SEQ ID NO:8). The fusion proteins were purified as described in Example 2A above. The fusion proteins were tested in a molecular beacon assay at various temperatures, as shown in FIGS. 3B-3C, FIG. 4 and FIG. 5 and their thermostability compared to variant RNA polymerases that were not fused. M13, M18 and M20 variants in FIGS. 3B-3C, FIG. 4 and FIG. 5 are examples of an amino acid sequence that is at least 80% sequence identity to SEQ ID NO:1; and include an amino acid substitution at positions corresponding to 109L, 205S, 388E, 534V, 567P and 618Q of SEQ ID NO:1.

[0076] The yields were compared using a molecular beacon assay that monitors in vitro transcription of a 6 Kb transcript. The in vitro transcription reactions were performed in 41 mM Tris-HCl pH 8.0, 50 mM NaCl, 19 mM MgCl.sub.2, 5.5 mM DTT, 1 mM spermidine, 4 mM of each ribonucleotide, 4.15 units/mL yeast inorganic pyrophosphatase, 1000 units/mL murine ribonuclease inhibitor, 30 nM DNA template, 30 nM RNA polymerase, and 0.5 .mu.M molecular beacon probe. A linearized plasmid DNA was used as template for the in vitro transcription reactions. The molecular beacon was designed to bind a 24 nucleotide target site upstream of the linearization site of the plasmid with a sequence of: 5'-CCT GC GATT GAA CAC GTG GGT CAG AGA GG GCAGG-3' (SEQ ID NO:9). The molecular beacons were labeled with the fluorescent dye TYE665 at the 5' end and the quencher IAbRQSp at the 3' end (or with the fluorescent dye 6-FAM at the 5' end and the quencher BHQ1 at the 3' end) (Integrated DNA Technologies, Coralville, Iowa). Reactions were run at various temperatures using a CFX96 Touch Real-Time PCT Detection System for one hour. The graph denotes end-point fluorescence units (representing the final yield from the in vitro transcription reaction) obtained for each polymerase plotted against the temperature at which the reactions were run.

Example 4: NASBA Assays

[0077] NASBA reactions were performed by first incubating RNA template (Jurkat total RNA), 250 nM forward primer (5'-AATTCTAATACGACTCACTATAGGGAGAGGCCCGGCATGTGGTGCATAA-3'; SEQ ID NO:71), 250 nM reverse primer (5'-CAGTATGCCAAGACCGACTCAGA-3'; SEQ ID NO:72) and 100 nM molecular beacon (5'-FAM-CGTACGAGAAGAGGAAGCCCAAGAGCCACGTACG-BHQ1-3', in which "FAM" refers to the dye 6-carboxyfluorescein and "BHQ" is Black Hole Quencher 1; SEQ ID NO:73) in 1.times.NASBA Buffer (40 mM Tris-HCl pH 8.5, 12 mM MgCl.sub.2, 70 mM KCl, 10 mM DTT) with 1 mM each dATP, dCTP, dGTP and dTTP, 2 mM each ATP, CTP, GTP, and UTP, and 0-5% v/v DMSO as indicated. Water was added in place of RNA to the -RNA control reactions. RNA+primer mixture was heated to 65.degree. C. for 2 minutes and temperature reduced to 40-56.degree. C. as indicated. After 10-minute incubation at secondary temperature, NASBA enzymes were added at final concentrations of 2-10 ng/.mu.L RNA polymerase variant (M20), 50-250 ng/.mu.L Reverse Transcriptase variant, and 0.005-0.02 U/.mu.L Thermus thermophilus RNase H. Incubation continued at the indicated gradient (40-56.degree. C.) temperatures and data was collected by fluorescence measurement every 30 seconds using the FAM channel of a Bio-Rad CFX96 real time instrument. Instrument-defined threshold times are shown in FIGS. 7A and 7B, with positive (+RNA) reactions indicated by squares and solid line, negative (-RNA) reactions by triangles and dashed line. This data shows that NASBA reactions that are done at a higher temperature (e.g., in the range of 45.degree. C. to 60.degree. C.) produce less non-specific amplification products relative to the same reactions that are done at a lower temperature (e.g., 41.degree. C.), and that the effect is not dependent upon DMSO.

Sequence CWU 1

1

771883PRTArtificial SequenceSynthetic construct 1Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe865 870 875 880Ala Phe Ala218DNAArtificial SequenceSynthetic construct 2taatacgact cactatag 18318DNAArtificial SequenceSynthetic construct 3aattaaccct cactaaag 18418DNAArtificial SequenceSynthetic construct 4tatttaccct cactaaag 18523DNAArtificial SequenceSynthetic constructmisc_feature(22)..(22)n is a, c, g, or t 5atttaggtga cactatagaa gng 23663PRTArtificial SequenceSynthetic construct 6Ala Thr Val Lys Phe Lys Tyr Lys Gly Glu Glu Lys Glu Val Asp Ile1 5 10 15Ser Lys Ile Lys Lys Val Trp Arg Val Gly Lys Met Ile Ser Phe Thr 20 25 30Tyr Asp Glu Gly Gly Asp Lys Thr Gly Arg Gly Ala Val Ser Glu Lys 35 40 45Asp Ala Pro Lys Glu Leu Leu Gln Met Leu Glu Lys Gln Lys Lys 50 55 60788PRTPyrococcus furiosus 7Gly Arg Lys Val Arg Thr Gln Gln Asn Glu Ile Leu Asn Leu Leu Asn1 5 10 15Glu Lys Glu Lys Ala Val Leu Arg Ala Ile Leu Glu His Gly Gly Glu 20 25 30Ile Lys Gln Glu Asp Leu Pro Glu Leu Val Gly Tyr Ser Arg Pro Thr 35 40 45Ile Ser Lys Val Ile Gln Glu Leu Glu Asn Lys Gly Leu Ile Lys Arg 50 55 60Glu Lys Ser Gly Lys Thr Phe Val Val Lys Ile Glu Arg Lys Ile Lys65 70 75 80Leu Asp Lys Met Gly Ala Pro Thr 85860PRTThermotoga maritima 8Lys Arg Arg Pro Thr Ile Asn Asp Val Ala Lys Leu Ala Gly Val Ser1 5 10 15Ile Ser Thr Val Ser Arg Tyr Leu Lys Asp Pro Ser Gln Val Ser Glu 20 25 30Lys Leu Gly Glu Arg Ile Arg Glu Ala Ile Lys Lys Leu Gly Tyr Lys 35 40 45Pro Asn Lys Ile Ala Gln Gly Leu Arg Thr Gly Asp 50 55 60934DNAArtificial SequenceSynthetic construct 9cctgcgattg aacacgtggg tcagagaggg cagg 3410142PRTPyrococcus furiosus 10Met Lys Thr Phe Leu Thr Glu Gln Gln Ile Lys Val Leu Met Leu Arg1 5 10 15Ala Lys Gly Tyr Lys Gln Ser Glu Ile Ala Lys Ile Leu Gly Thr Ser 20 25 30Arg Ala Asn Val Ser Ile Leu Glu Lys Arg Ala Met Glu Lys Ile Glu 35 40 45Lys Ala Arg Asn Thr Leu Leu Leu Trp Glu Gln Ile Asn Ser Lys Val 50 55 60Ile Val Glu Ile Lys Ala Gly Glu Asp Ile Phe Ser Ile Pro Glu Lys65 70 75 80Phe Phe Lys Lys Ala Asp Lys Val Gly Val Lys Val Pro Tyr Ser Thr 85 90 95Ala Glu Ile Ile Thr Phe Leu Val Glu His Ala Pro Val Glu Asp Arg 100 105 110Leu Ala Lys Arg Asp Phe Val Leu Phe Leu Asp Ser Lys Asn Lys Leu 115 120 125Arg Ile Gly Asp Cys Leu Val Ile Glu Glu Ile Lys Glu Asp 130 135 1401175PRTPyrococcus furiosus 11Met Pro Ile Thr Lys Val Thr Arg Asn Tyr Gln Ile Thr Ile Pro Ala1 5 10 15Glu Ile Arg Lys Ala Leu Gly Ile Lys Glu Gly Glu Leu Leu Glu Val 20 25 30Arg Leu Glu Asn Gly Lys Ile Ile Ile Glu Arg Leu Lys Lys Glu Arg 35 40 45Lys Thr Leu Lys Leu Gly Lys Lys Leu Thr Leu Glu Glu Ile Glu Lys 50 55 60Ala Ile Glu Glu Gly Met Lys Gln Cys Met Gln65 70 751276PRTPyrococcus furiosus 12Thr Lys Ile Glu Ile Leu Arg Leu Leu Lys Glu Arg Glu Met Tyr Ala1 5 10 15Tyr Glu Ile Trp Ser Leu Leu Gly Lys Pro Leu Lys Tyr Gln Ala Val 20 25 30His Gln His Ile Lys Glu Leu Leu Glu Leu Gly Leu Val Glu Gln Ala 35 40 45Tyr Arg Lys Gly Lys Arg Val Tyr Tyr Lys Ile Thr Glu Lys Gly Leu 50 55 60Arg Ile Leu Gln Asn Phe Glu Asp Leu Glu Asn Ile65 70 751372PRTPyrococcus furiosus 13Met Asn Thr Gly Ala Gln Gly Val Ser Glu Met Ser Arg Met Lys Ile1 5 10 15Ile Ser Val Gln Leu Pro Gln Ser Leu Ile His Gly Leu Asp Ala Leu 20 25 30Val Lys Arg Gly Ile Tyr Pro Asn Arg Ser Glu Ala Ile Arg Val Ala 35 40 45Ile Arg Glu Leu Leu Lys Lys Glu Leu Tyr Lys Glu Glu Ile Gln Glu 50 55 60Glu Ile Pro Glu Tyr Val Val Lys65 701467PRTPyrococcus furiosus 14Val Ile Ile Pro Arg Pro Ile Asp Pro Arg Asp Ile Arg Arg Ile Arg1 5 10 15Lys Glu Leu Gly Ile Thr Gln Glu Glu Leu Ala Arg Lys Ala Gly Val 20 25 30Thr Gln Ala Tyr Ile Ala Lys Leu Glu Ala Gly Lys Val Asp Pro Arg 35 40 45Leu Ser Thr Phe Asn Lys Ile Leu Arg Ala Leu Ile Glu Cys Gln Lys 50 55 60Ala Lys Ile651565PRTPyrococcus furiosus 15Asn Asn Cys Glu Cys Met Val Val Lys Glu Lys Leu Tyr Thr Val Lys1 5 10 15Gln Ala Ser Glu Ile Leu Gly Val His Pro Lys Thr Ile Gln Lys Trp 20 25 30Asp Arg Glu Gly Lys Ile Lys Thr Val Arg Thr Pro Gly Gly Arg Arg 35 40 45Arg Ile Pro Glu Ser Glu Ile Lys Arg Leu Leu Gly Ile Ser Glu Glu 50 55 60Lys651671PRTPyrococcus furiosus 16Met Leu Lys Asp Ser Ala Pro Lys Arg Lys Ile Leu Glu Glu Leu Arg1 5 10 15Lys Gly Glu Thr Val Ser Gly Asp Tyr Leu Ala Ser Lys Leu Gly Val 20 25 30Ser Arg Val Ala Ile Trp Lys His Ile Arg Glu Leu Lys Glu Leu Gly 35 40 45Tyr Gly Ile Ile Ala Asp Lys Lys Gly Tyr Lys Leu Val Tyr Glu Pro 50 55 60Lys Lys Pro Tyr Pro Trp Glu65 701786PRTPyrococcus furiosus 17Met Ile Asp Glu Arg Asp Lys Ile Ile Leu Glu Ile Leu Glu Lys Asp1 5 10 15Ala Arg Thr Pro Phe Thr Glu Ile Ala Lys Lys Leu Gly Ile Ser Glu 20 25 30Thr Ala Val Arg Lys Arg Val Lys Ala Leu Glu Glu Lys Gly Ile Ile 35 40 45Glu Gly Tyr Thr Ile Lys Ile Asn Pro Lys Lys Leu Gly Tyr Ser Leu 50 55 60Val Thr Ile Thr Gly Val Asp Thr Lys Pro Glu Lys Leu Phe Glu Val65 70 75 80Ala Glu Lys Leu Lys Glu 851877PRTPyrococcus furiosus 18Met Glu Ile Asp Asp Leu Asp Arg Lys Ile Leu Ser Leu Leu Ile Glu1 5 10 15Asp Ser Arg Leu Ser Tyr Arg Glu Ile Ala Lys Lys Leu Asn Val Ala 20 25 30Val Gly Thr Ile Tyr Asn Arg Ile Lys Lys Leu Glu Asp Met Gly Val 35 40 45Ile Gln Gly Phe Thr Val Lys Leu Asn Tyr Glu Lys Leu Gly Tyr Glu 50 55 60Leu Thr Ala Ile Ile Gly Ile Lys Ala Gln Gly Lys Lys65 70 751994PRTPyrococcus furiosus 19Glu Met Leu Trp Met Tyr Ile Leu Lys Leu Leu Lys Asp Arg Pro Met1 5 10 15Tyr Ala Tyr Glu Ile Arg Asn Glu Leu Lys Lys Arg Phe Gly Phe Glu 20 25 30Pro Ala Thr Val Ser Ser Tyr Val Val Leu Tyr Arg Leu Glu Glu Gly 35 40 45Gly Tyr Val Ser Ser Glu Trp His Glu Ser Glu Ala Gly Arg Pro Ser 50 55 60Arg Lys Tyr Tyr Arg Leu Thr Glu Lys Gly Glu Lys Leu Leu Glu Lys65 70 75 80Gly Ile Glu Thr Ile Glu Asp Val Leu Asn Met Leu Lys Ser 85 902068PRTPyrococcus furiosus 20Met Lys Val Ser Lys Ala Thr Ala Ser Lys Val Leu Arg Ser Leu Glu1 5 10 15Asn Lys Gly Ile Val Glu Arg Glu Arg Arg Gly Lys Thr Tyr Leu Val 20 25 30Arg Leu Thr Asn Lys Gly Leu Glu Leu Leu Glu Glu Ile Ser Lys Ala 35 40 45Gly Lys Glu Leu Asp Glu Lys Ile Phe Ala Glu Met Ser Val Asp Glu 50 55 60Arg Ile Val Leu652156PRTPyrococcus furiosus 21Ser Glu Asp Tyr Met Leu Gln Asn Arg Arg Lys Val Leu Ala Lys Val1 5 10 15Leu Glu Leu Leu Asn Tyr Asn Pro Lys Ala Leu Asn Ile Ser Glu Leu 20 25 30Ala Arg Met Phe Gly Val Ser Arg Asp Thr Ile Tyr Asn Asp Ile Gln 35 40 45Gln Ile Ile Lys Asn Val Glu Val 50 552294PRTPyrococcus furiosus 22Ser Lys Glu Ile Ser Arg Phe Leu Lys Val Ile Ser Asn Pro Ile Arg1 5 10 15Tyr Gly Ile Leu Lys Met Leu Asn Asp Arg Trp Met Cys Val Cys Leu 20 25 30Ile Ser Glu Ala Leu Glu Ile Asp Gln Thr Leu Val Ser His His Ile 35 40 45Arg Ile Leu Lys Glu Leu Asp Leu Leu Glu Glu Arg Lys Glu Gly Lys 50 55 60Leu Arg Phe Tyr Arg Thr Asn Lys Glu Lys Leu Arg Glu Tyr Leu Glu65 70 75 80Lys Val Leu Glu Asp Phe Asn His Gly Thr Ser Lys Gly Ser 85 902378PRTPyrococcus furiosus 23Met Cys Arg Lys Asp Val Met Ile Ile Ser Asp Pro Lys Gln Ile Lys1 5 10 15Ala Leu Ser Asp Pro Thr Arg Val Lys Ile Leu Glu Leu Leu Arg Tyr 20 25 30His Pro Met Thr Val Ser Glu Ile Ser Arg Val Ile Gly Lys Asp Lys 35 40 45Ser Thr Ile Tyr Arg His Ile Lys Ala Leu Glu Glu Ala Gly Leu Val 50 55 60Glu Glu Val Glu Lys Ile Gly Asn Glu Thr Val Tyr Gly Arg65 70 7524122PRTPyrococcus furiosus 24Met Glu Pro Val Glu Phe Lys Leu Asn Gln Lys Gly Ile Lys Ser Ile1 5 10 15Leu Pro Thr Met Glu Ala Glu Ile Met Glu Tyr Met Trp Glu Ile Lys 20 25 30Glu Ala Thr Ala Gly Glu Val Tyr Glu Tyr Met Lys Thr Lys Tyr Pro 35 40 45Glu Ile Arg Arg Ser Thr Val Ser Ile Leu Met Asn Arg Leu Cys Glu 50 55 60Arg Gly Leu Leu Lys Arg Arg Met Glu Lys Gly Lys Gly Gly Ile Arg65 70 75 80Tyr Val Tyr Ser Ile Thr Thr Thr Arg Glu Glu Phe Glu Arg Lys Val 85 90 95Val Glu Lys Ile Ile Glu Ser Leu Met Met Asn Phe Arg Glu Ala Thr 100 105 110Phe Ala Tyr Leu Ser

Lys Ile Asn Lys Lys 115 1202587PRTPyrococcus furiosus 25Met Lys Lys Ser Asn Leu Asp Leu Leu Ile Leu Leu Ala Lys Ala Gly1 5 10 15Gly Ile Glu Lys Glu Ile Leu Thr Thr Ser Arg Glu Leu Ser Lys Met 20 25 30Leu Asn Val Ser Pro Gln Thr Ile Val Arg Trp Leu Glu Asp Leu Glu 35 40 45Lys Asp Gly Leu Ile Lys Lys Ser Glu Ser Arg Lys Gly Thr Leu Val 50 55 60Thr Ile Thr Glu Glu Gly Val Lys Phe Leu Glu Lys Leu His Glu Glu65 70 75 80Leu Ser Asp Ala Leu Tyr Arg 8526104PRTUnknownThermococcaceae 26Met Glu Ile Pro Pro Glu Ile Ser His Ala Leu Ser Glu Ile Gly Phe1 5 10 15Thr Lys Tyr Glu Ile Leu Thr Tyr Trp Thr Leu Leu Val Tyr Gly Pro 20 25 30Ser Thr Ala Lys Glu Ile Ser Thr Lys Ser Gly Ile Pro Tyr Asn Arg 35 40 45Val Tyr Asp Thr Ile Ser Ser Leu Lys Leu Arg Gly Phe Val Thr Glu 50 55 60Ile Glu Gly Thr Pro Lys Val Tyr Ala Ala Tyr Ser Pro Arg Ile Ala65 70 75 80Phe Phe Arg Phe Lys Lys Glu Leu Glu Asp Ile Met Lys Lys Leu Glu 85 90 95Ile Glu Leu Asn Asn Val Lys Lys 1002775PRTPyrococcus furiosus 27Ile Ile Asn Pro Gln Ala Arg Leu Thr Pro Leu Glu Leu Glu Ile Leu1 5 10 15Glu Ile Ile Lys Gln Lys Lys Ser Ile Thr Ile Thr Glu Ile Lys Glu 20 25 30Ile Leu Ser Glu Arg Arg Lys Ser Glu Tyr Pro Leu Ser Leu Val Ser 35 40 45Glu Tyr Ile Ser Arg Leu Glu Arg Lys Gly Tyr Val Lys Lys Ile Ala 50 55 60Lys Gly Arg Lys Lys Phe Val Glu Ala Leu Ile65 70 752894PRTPyrococcus furiosus 28Gly Ile Asp Val Val Ile Pro Glu Ile Lys His Asp Pro Ile Ala Arg1 5 10 15Asp Ile Val Lys Ile Leu Phe Asp Leu Arg Arg Ala Asn Val Ser Gln 20 25 30Ile Ala Arg Glu Leu Lys Gly Arg Arg Gly Lys Ala Ser Arg Asn Thr 35 40 45Val Arg Lys Lys Leu Lys Glu Leu Glu Lys Leu Gly Val Val Lys Glu 50 55 60Val Pro Gly Glu Arg Gly Ser Val Tyr Thr Leu Ser Arg Glu Val Val65 70 75 80Lys Lys Trp Leu Asp Leu Ile Gly Ile Pro Ile Asn Leu Leu 85 902967PRTPyrococcus furiosus 29Met Thr Lys Arg Val Lys Val Ile Thr Asp Pro Glu Val Ile Lys Val1 5 10 15Met Leu Glu Asp Thr Arg Arg Lys Ile Leu Gln Leu Leu Arg Asn Arg 20 25 30Glu Met Thr Ile Ser Gln Leu Ser Glu Ile Leu Gly Lys Met Pro Gln 35 40 45Thr Ile Tyr His His Ile Glu Lys Leu Lys Glu Ala Gly Leu Val Glu 50 55 60Val Lys Arg6530100PRTPyrococcus furiosus 30Met Glu Glu Ile Lys Glu Ile Met Lys Ser His Thr Leu Gly Asn Pro1 5 10 15Val Arg Leu Gly Ile Met Ile Tyr Leu Phe Pro Arg Arg Arg Ala Pro 20 25 30Phe Ser His Ile Gln Lys Ala Leu Asp Leu Thr Pro Gly Asn Leu Asp 35 40 45Ser His Ile Lys Val Leu Glu Lys His Gly Phe Val Arg Thr Tyr Lys 50 55 60Val Ile Ala Asp Arg Pro Arg Thr Met Val Glu Ile Thr Asp Tyr Gly65 70 75 80Met Glu Glu Thr Arg Lys Phe Leu Ser His Leu Lys Thr Val Ile Asp 85 90 95Ala Ile His Phe 1003199PRTPyrococcus furiosus 31Met Gly Glu Glu Leu Asn Arg Leu Leu Asp Val Leu Gly Asn Glu Thr1 5 10 15Arg Arg Arg Ile Leu Phe Leu Leu Thr Lys Arg Pro Tyr Phe Val Ser 20 25 30Glu Leu Ser Arg Glu Leu Gly Val Gly Gln Lys Ala Val Leu Glu His 35 40 45Leu Arg Ile Leu Glu Glu Ala Gly Leu Ile Glu Ser Arg Val Glu Lys 50 55 60Ile Pro Arg Gly Arg Pro Arg Lys Tyr Tyr Met Ile Lys Lys Gly Leu65 70 75 80Arg Leu Glu Ile Leu Leu Thr Pro Thr Leu Phe Gly Ser Glu Met Tyr 85 90 95Glu Ala Lys3261PRTPyrococcus furiosus 32Met Arg Arg Met Asp Lys Val Asp Leu Gln Leu Ile Lys Ile Leu Ser1 5 10 15Gln Asn Ser Arg Leu Thr Tyr Arg Glu Leu Ala Glu Met Leu Gly Thr 20 25 30Thr Arg Gln Arg Val Ala Arg Lys Val Asp Lys Leu Lys Lys Leu Gly 35 40 45Ile Ile Arg Lys Phe Thr Ile Ile Pro Asn Leu Glu Lys 50 55 603382PRTPyrococcus furiosus 33Gly Arg Lys Val Arg Thr Gln Gln Asn Glu Ile Leu Asn Leu Leu Asn1 5 10 15Glu Lys Glu Lys Ala Val Leu Arg Ala Ile Leu Glu His Gly Gly Glu 20 25 30Ile Lys Gln Glu Asp Leu Pro Glu Leu Val Gly Tyr Ser Arg Pro Thr 35 40 45Ile Ser Lys Val Ile Gln Glu Leu Glu Asn Lys Gly Leu Ile Lys Arg 50 55 60Glu Lys Ser Gly Lys Thr Phe Val Val Lys Ile Glu Arg Lys Ile Lys65 70 75 80Leu Asp3459PRTThermotoga maritima 34Lys Ser Leu Gln Arg Phe Leu Arg Arg Asn Thr Thr Ser Ile Lys His1 5 10 15Leu Ser Glu Ile Thr Gly Val Ala Arg Asn Arg Leu Ser Asp Ile Leu 20 25 30Asn Gly Lys Thr Gln Lys Ile Arg Gly Glu Thr Leu Arg Lys Ile Ala 35 40 45Lys Ala Phe Glu Lys Ser Asn Ile Leu Ser Phe 50 5535107PRTThermotoga 35Asp Val Ile Gln Arg Ile Lys Glu Lys Tyr Asp Glu Phe Thr Asn Ala1 5 10 15Glu Lys Lys Ile Ala Asp Thr Ile Leu Ser Asp Pro Lys Gly Ile Ile 20 25 30Glu Ser Ser Ile Ser Asp Leu Ser Glu Lys Ala Gly Val Lys Ser Glu 35 40 45Ala Ser Val Val Lys Phe Tyr Lys Lys Leu Gly Leu Asn Ser Phe Gln 50 55 60Gln Phe Lys Val Leu Leu Ala Gln Ser Ile Ser Arg Ala Pro Leu Glu65 70 75 80Ile Val Tyr Glu Asp Val Ser Ser Glu Asp Asp Thr Lys Thr Ile Thr 85 90 95Glu Lys Ile Phe Lys Ala Thr Val Arg Ala Ile 100 10536101PRTThermotoga maritima 36Lys Ile Arg Asp Lys Ile Leu Asn Val Tyr Thr Gln Phe Ser Pro Ala1 5 10 15Glu Arg Lys Val Ala Asp Tyr Val Leu Glu Arg Pro Asp Asp Val Ile 20 25 30His Tyr Ser Ile Thr Glu Phe Ala Lys Ile Val Gly Val Ser Glu Thr 35 40 45Thr Ile His Arg Met Ile Lys Lys Leu Asp Phe Glu Gly Tyr Gln Ala 50 55 60Phe Lys Ile Ala Leu Ala Arg Glu Leu Ser Gly Leu Glu Glu Thr Ile65 70 75 80Glu Arg Arg Asp Phe Ile Asp Glu Glu Ile Asp Ile Leu Arg Arg Leu 85 90 95Lys Asp Thr Leu Asp 1003760PRTThermotoga maritima 37Lys Arg Arg Pro Thr Ile Asn Asp Val Ala Lys Leu Ala Gly Val Ser1 5 10 15Ile Ser Thr Val Ser Arg Tyr Leu Lys Asp Pro Ser Gln Val Ser Glu 20 25 30Lys Leu Gly Glu Arg Ile Arg Glu Ala Ile Lys Lys Leu Gly Tyr Lys 35 40 45Pro Asn Lys Ile Ala Gln Gly Leu Arg Thr Gly Asp 50 55 603858PRTThermotoga maritima 38Met Ala Ser Ile Lys Asp Val Ala Lys Leu Ala Gly Val Ser Ile Ala1 5 10 15Thr Val Ser Arg Val Ile Asn Gly Tyr Asn Asn Val Ser Glu Glu Thr 20 25 30Arg Lys Lys Val Ile Asp Ala Ile Arg Lys Leu Asn Tyr His Pro Val 35 40 45Tyr Ala Val Lys Gly Ala Val Leu Lys Arg 50 553961PRTThermotoga maritima 39Met Lys Lys Lys Tyr Val Thr Ile Arg Asp Ile Ala Glu Lys Ala Gly1 5 10 15Val Ser Ile Asn Thr Val Ser Arg Ala Leu Asn Asn Lys Pro Asp Ile 20 25 30Ser Glu Glu Thr Arg Arg Lys Ile Leu Lys Ile Ala Gln Glu Leu Gly 35 40 45Tyr Val Lys Asn Ala Thr Ala Ser Ser Leu Arg Ser Lys 50 55 604058PRTThermotoga 40Met Pro Thr Ile Glu Asp Val Ala Lys Leu Ala Gly Val Ser Ile Ala1 5 10 15Thr Val Ser Arg Val Ile Asn Gly Ser Gly Tyr Val Ser Glu Lys Thr 20 25 30Arg Tyr Lys Val Trp Lys Ala Ile Glu Glu Leu Gly Tyr Lys Pro Glu 35 40 45Ile Ser Ala Lys Leu Leu Ala Ser Lys Gly 50 554161PRTThermotoga maritima 41Met Arg Ile Gly Glu Lys Leu Arg Lys Leu Arg Leu Ser Arg Gly Leu1 5 10 15Thr Gln Glu Glu Leu Ala Glu Arg Thr Asp Leu Ser Arg Ser Phe Ile 20 25 30Ser Gln Leu Glu Ser Asp Lys Thr Ser Pro Ser Ile Asp Thr Leu Glu 35 40 45Arg Ile Leu Glu Ala Leu Gly Thr Asp Leu Lys His Phe 50 55 604267PRTThermotoga maritima 42Met His Met Lys Thr Val Arg Gln Glu Arg Leu Lys Ser Ile Val Arg1 5 10 15Ile Leu Glu Arg Ser Lys Glu Pro Val Ser Gly Ala Gln Leu Ala Glu 20 25 30Glu Leu Ser Val Ser Arg Gln Val Ile Val Gln Asp Ile Ala Tyr Leu 35 40 45Arg Ser Leu Gly Tyr Asn Ile Val Ala Thr Pro Arg Gly Tyr Val Leu 50 55 60Ala Gly Gly654375PRTThermotoga maritima 43Met Asn Thr Leu Lys Lys Ala Phe Glu Ile Leu Asp Phe Ile Val Lys1 5 10 15Asn Pro Gly Asp Val Ser Val Ser Glu Ile Ala Glu Lys Phe Asn Met 20 25 30Ser Val Ser Asn Ala Tyr Lys Tyr Met Val Val Leu Glu Glu Lys Gly 35 40 45Phe Val Leu Arg Lys Lys Asp Lys Arg Tyr Val Pro Gly Tyr Lys Leu 50 55 60Ile Glu Tyr Gly Ser Phe Val Leu Arg Arg Phe65 70 754481PRTThermotoga maritima 44Met Lys Ile Ser Lys Lys Arg Arg Gln Glu Leu Ile Arg Lys Ile Ile1 5 10 15His Glu Lys Lys Ile Ser Asn Gln Phe Gln Ile Val Glu Glu Leu Lys 20 25 30Lys Tyr Gly Ile Lys Ala Val Gln Pro Thr Val Ala Arg Asp Leu Lys 35 40 45Glu Ile Gly Ala Val Lys Ile Met Asp Glu Ser Gly Asn Tyr Val Tyr 50 55 60Lys Leu Leu Asp Glu Thr Pro Val Ile Asp Pro Trp Lys Glu Leu Lys65 70 75 80Arg4582PRTThermotoga maritima 45Met His Lys Lys Leu Asn Pro Lys Ser Met Lys Arg Glu Asn Lys Lys1 5 10 15Met Val Leu Arg Tyr Leu Ile Glu Ser Gly Pro His Ser Arg Val Glu 20 25 30Ile Ala Arg Lys Thr Gly Leu Ala Gln Ser Ala Ile Trp Arg Ile Ile 35 40 45Glu Glu Leu Val Asn Glu Gly Leu Val Glu Glu Lys Gly Thr Ala Thr 50 55 60Gly Arg Arg Arg Lys Ala Val Thr Tyr Gly Pro Thr Arg Ser Phe Ile65 70 75 80Thr Ser4679PRTThermotoga maritima 46Met Pro Ser Pro Leu Leu Arg Arg Glu Asn Lys Ile Lys Ile Leu Arg1 5 10 15Tyr Ile Leu Lys Asn Gly Lys Thr Thr Arg Asn Gln Leu Ala Ser Asn 20 25 30Leu Asn Leu Ala His Ser Thr Leu Ser Tyr Ile Ile Asp Glu Leu Leu 35 40 45Asp Glu Gly Phe Leu Val Phe Glu Glu Ile Lys Lys Lys Arg Gly Arg 50 55 60Pro Tyr Gln Ile Leu Ser Val Asn Pro Glu Lys Phe Thr Ala Ile65 70 754779PRTThermotoga maritima 47Met Lys Glu Glu Arg Leu Lys Glu Ile Leu Asp Ile Val Asp Arg Asn1 5 10 15Gly Phe Ile Ser Met Lys Asp Leu Gln Glu Gln Leu Gly Val Ser Met 20 25 30Ile Thr Val Arg Arg Asp Val Ala Glu Leu Val Lys Arg Asn Leu Val 35 40 45Lys Lys Val His Gly Gly Ile Arg Lys Val Asn Tyr Phe Glu Lys Glu 50 55 60Thr Asp Phe Met Lys Arg Leu Ser Ile Asn Arg Glu Ala Lys Glu65 70 7548137PRTThermotoga maritima 48Met Phe Thr Met Arg Ser Glu Tyr Ala Leu Arg Leu Met Ile Val Met1 5 10 15Ala Lys Glu Tyr Gly Asn Tyr Leu Ser Met Thr Glu Ile Leu Glu Lys 20 25 30Ala Lys Gln Ser Val Pro Arg Glu Phe Ala Glu Lys Ile Leu Tyr Thr 35 40 45Leu Lys Lys Ala Gly Leu Val Lys Thr Arg Arg Gly Lys Ser Gly Gly 50 55 60Tyr Met Leu Ser Arg Pro Pro Lys Glu Ile Lys Val Ser Glu Ile Val65 70 75 80Phe Leu Leu Asp Arg Lys Ser Lys Val Phe Phe Asp Met Pro Gly Cys 85 90 95Pro Asp Glu Leu Asp Cys Val Ile Arg Ala Leu Trp Lys Arg Val Glu 100 105 110Asn Glu Ile Glu Lys Ile Leu Ser Gly Val Thr Leu Glu Asp Leu Val 115 120 125Arg Glu Gln Glu Glu Lys Met Lys Gln 130 1354995PRTThermotoga naphthophila 49Met Arg Asp Thr Lys Gly His Leu Lys Phe Leu Val Leu His Ile Ile1 5 10 15Ser Gln Gln Pro Ser His Gly Tyr Tyr Ile Met Lys Lys Ile Ser Gln 20 25 30Ile Ile Gly Ala Glu Pro Pro Ser Pro Gly Ala Leu Tyr Pro Ile Leu 35 40 45Ser Ser Leu Arg Lys Gln Lys Tyr Ile Glu Thr Tyr Asn Glu Gly Lys 50 55 60Arg Lys Val Tyr Arg Leu Thr Asp Lys Gly Arg Lys Tyr Leu Glu Glu65 70 75 80His Lys Glu Glu Ile Lys Lys Ala Leu Asp Phe Ala Glu Arg Phe 85 90 9550118PRTThermotoga maritima 50Met Arg His Arg Gly Gly Arg Gly Phe Arg Gly Trp Trp Leu Ala Ser1 5 10 15Thr Ile Leu Leu Leu Val Ala Glu Lys Pro Ser His Gly Tyr Glu Leu 20 25 30Ala Glu Arg Leu Ala Glu Phe Gly Ile Glu Ile Pro Gly Ile Gly His 35 40 45Met Gly Asn Ile Tyr Arg Val Leu Ala Asp Leu Glu Glu Ser Gly Phe 50 55 60Leu Ser Thr Glu Trp Asp Thr Thr Val Ser Pro Pro Arg Lys Ile Tyr65 70 75 80Arg Ile Thr Pro Gln Gly Lys Leu Tyr Leu Arg Glu Ile Leu Arg Ser 85 90 95Leu Glu Asp Met Lys Arg Arg Ile Glu Thr Leu Glu Glu Arg Ile Lys 100 105 110Arg Val Leu Gln Glu Glu 1155170PRTThermotoga maritima 51Met Leu Ser Lys Arg Asp Ala Ile Leu Lys Ala Ala Val Glu Val Phe1 5 10 15Gly Lys Lys Gly Tyr Asp Arg Ala Thr Thr Asp Glu Ile Ala Glu Lys 20 25 30Ala Gly Val Ala Lys Gly Leu Ile Phe His Tyr Phe Lys Asn Lys Glu 35 40 45Glu Leu Tyr Tyr Gln Ala Tyr Met Ser Val Thr Glu Lys Leu Gln Lys 50 55 60Glu Phe Glu Asn Phe Leu65 7052883PRTEnterobacteria bacteriophage 13a 52Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Val Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn

Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Asp Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Leu Asp Glu Ile Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Val Ile Leu Gln Ala Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala53883PRTYersinia bacteriophage YpP-R 53Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Asn Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Thr Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Leu Asp Glu Val Gly Gly Leu Ala Val Asn545 550 555 560Leu Leu Pro Ser Ala Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Val Ile Leu Gln Ala Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala54883PRTYersinia bacteriophage R 54Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Asn Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Thr Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Leu Asp Glu Val Gly Gly Leu Ala Val Asn545 550 555 560Leu Leu Pro Ser Ala Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Val Ile Leu Gln Ala Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Pro Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp

Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala55883PRTYersinia bacteriophage phiA1122 55Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Asn Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Thr Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Leu Asp Glu Val Gly Gly Leu Ala Val Asn545 550 555 560Leu Leu Pro Ser Ala Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Val Ile Leu Gln Ala Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Pro Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala56883PRTEscherichia bacteriophage CICC 80001 56Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Val Val Lys Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ser Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Arg Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Met Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ser Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Leu Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Val Ile Leu Gln Glu Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Ile Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala57903PRTYersinia bacteriophage YpsP-G 57Met Thr Glu Arg Thr Asp Gly Leu Lys Lys Gly Tyr Met Pro Asn Gly1 5 10 15Thr Leu Tyr Ala Ala Asn Arg Arg Leu Val Arg Thr Trp Arg Glu Asn 20 25 30Asn Leu Glu Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr 35 40 45Gly Glu Arg Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr 50 55 60Glu Met Gly Glu Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys65 70 75 80Ala Gly Glu Val Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr 85 90 95Leu Leu Pro Lys Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val 100 105 110Lys Ala Lys Arg Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu 115 120 125Ile Lys Pro Glu Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala 130 135 140Cys Leu Thr Ser Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala145 150 155 160Ile Gly Arg Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp 165 170 175Leu Glu Ala Lys His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys 180 185 190Arg Val Gly His Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala 195 200 205Asp Met Leu Ser Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp 210 215 220His Lys Glu Asp Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu225 230 235 240Ile Glu Ser Thr Gly Met Val Asn Leu His Arg Gln Asn Ala Gly Val 245 250 255Val Gly Gln Asp Ser Glu Thr Ile Glu Leu Thr Pro Glu Tyr Ala Glu 260 265 270Ala Ile Ala Thr Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe 275 280 285Gln Pro Cys Val Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly 290 295 300Gly Tyr Trp Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His305 310 315 320Ser Lys Lys Ala Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val 325 330 335Tyr Lys Ala Ile Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys 340 345 350Lys Val Leu Ala Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro 355 360 365Val Glu Asp Ile Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro 370 375 380Glu Asp Ile Asp Thr Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala385 390 395 400Ala Ala Ala Val Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile 405 410 415Ser Leu Glu Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys 420 425 430Ala Ile Trp Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala 435 440 445Val Ser Met Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu 450 455 460Thr Leu Ala Lys Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu465 470 475 480Lys Ile His Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro 485 490 495Glu Arg Ile Lys Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys 500 505 510Ala Lys Ser Pro Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro 515 520 525Phe Cys Phe Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His 530 535 540Gly Leu Ser Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys545 550 555 560Ser Gly Ile Gln His Phe Ser Ala Met Leu Leu Asp Glu Val Gly Gly 565 570 575Leu Ala Val Asn Leu Leu Pro Ser Ala Thr Val Gln Asp Ile Tyr Gly 580 585 590Ile Val Ala Lys Lys Val Asn Val Ile Leu Gln Ala Asp Val Ile Asn 595 600 605Gly Thr Asp Asn Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu 610 615 620Ile Ser Glu Lys Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp625 630 635 640Leu Ala Tyr Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr 645 650 655Leu Ala Tyr Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu 660 665 670Asp Thr Ile Gln Pro Val Ile Asp Ser Gly Lys Gly Leu Met Phe Thr 675

680 685Gln Pro Asn Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala 690 695 700Val Ser Val Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys705 710 715 720Ser Ala Ala Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly 725 730 735Glu Ile Leu Arg Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly 740 745 750Phe Pro Val Trp Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn 755 760 765Leu Met Phe Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn 770 775 780Lys Asp Ser Glu Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro785 790 795 800Asn Phe Val His Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val 805 810 815Trp Ala His Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp 820 825 830Ser Phe Gly Thr Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val 835 840 845Arg Glu Thr Met Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp 850 855 860Phe Tyr Asp Gln Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys865 870 875 880Met Pro Ala Leu Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu 885 890 895Lys Ser Asp Phe Ala Phe Ala 90058883PRTSalmonella bacteriophage Vi06 58Met Asn Thr Ile Ser Ile Thr Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Val Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Ile 50 55 60Ala Asp Asn Asp Ala Thr Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Met Ile Ala Arg Ile Asn Ser Trp Phe Lys Glu Val Gln Ala Lys Cys 85 90 95Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Gly Ile Lys Pro Glu 100 105 110Ala Ile Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Arg Leu Thr Ser 115 120 125Met Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Ile Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ser Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Ser Ile Ser Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Ala Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Val305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Arg Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Val Val Thr Lys Trp Lys His Cys Pro Val Asp Tyr Ile 340 345 350Pro Thr Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Pro Glu Ala Leu Ala Ser Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Met Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Arg Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Phe Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Leu Ala Cys Ala Lys Ser Pro 485 490 495Leu Glu Asn Thr Trp Trp Ser Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Gly Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Phe Gly Ile Gln 530 535 540His Phe Ser Val Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Glu Ile Leu Gln Val Asp Met Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Asp Lys Thr Gly Glu Ile Tyr Glu Lys 595 600 605Ile Lys Leu Gly Thr Lys Glu Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr His Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Val Glu Val Lys Asp Arg Lys Thr Gly Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Lys Lys Pro Val Gln Thr Arg Leu Asn Leu Ile Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Arg Asp Ser Glu 755 760 765Ile Asp Ala Tyr Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Asp Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Ala Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Ala Gln 835 840 845Phe Ala Asp Gln Leu His Lys Ser Gln Leu Asp Lys Met Pro Val Leu 850 855 860Pro Ser Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala59883PRTStenotrophomonas bacteriophage IME15 59Met Thr Val Ile Ala Ile Glu Lys Asn Asp Phe Ser Asp Val Glu Leu1 5 10 15Ala Val Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Lys Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ala Tyr Glu Met Gly Glu 35 40 45Ala Arg Phe Arg Lys Ile Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Val Ala Thr Leu Leu Pro Lys65 70 75 80Met Ile Glu Arg Ile His Ala Trp Phe Glu Glu Val Ser Ala Lys Arg 85 90 95Gly Lys Arg Pro Thr Ala Phe Lys Phe Leu Gln Glu Val Lys Pro Glu 100 105 110Ala Ile Ala Tyr Ile Thr Ile Lys Thr Val Leu Gly Thr Leu Thr Ser 115 120 125Ala Glu Gln Thr Thr Val Gln Ala Ala Ala Ser Ala Val Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Leu Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Met Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ala Thr 210 215 220Gly Leu Val Val Leu Glu Arg Gln Asn Ala Gly Val Val Gly Ala Asp225 230 235 240Ala Glu Thr Leu Ser Leu Ala Ser Glu Tyr Ala Asp Ala Ile Ala Thr 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Tyr Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Thr Val Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Gly Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Val305 310 315 320Asn Leu Ala Gln Ser Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Ala Asn Glu Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Val Lys Pro Asp Asp Ile Asp 355 360 365Glu Asn Pro Glu Ala Leu Thr Asn Trp Lys Arg Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Leu Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Ala Ile Gly Lys Glu Gly Phe Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu His Ile Met Ala Ser Ala Lys Asn Pro 485 490 495Leu Glu Tyr Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Met His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575Lys Val Asn Glu Ile Met Gln Arg Asp Val Ile Ser Gly Thr Asp Asp 580 585 590Glu Leu Val Thr Glu Thr Asp Lys Thr Thr Gly Glu Ile Thr Glu Lys 595 600 605Ala Val Leu Gly Thr Arg Thr Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Ala Asn Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Arg 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Ile Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ser Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Lys Trp Leu Gln Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Val Leu Arg705 710 715 720Asn Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Arg Lys Pro Leu Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Gly 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Gly Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asp Thr Tyr Glu Asn Cys Asp Val Leu Ala Asp Phe Tyr Glu Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Lys Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe865 870 875 880Ala Phe Ala60884PRTCitrobacter bacteriophage SH2 60Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Val Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ser Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Val Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Asn Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470

475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala61884PRTEnterobacter bacteriophage E-4 61Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Ser Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ser Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Asn Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Leu Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala62884PRTYersinia bacteriophage phiYe-F10 62Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Val Ala Tyr Ala Pro Leu Gln Ser Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ser Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Ile Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Val Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala63884PRTCitrobacter bacteriophage phiCFP-1 63Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Asp Ser Lys 85 90 95Lys Gly Arg Lys Pro Val Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ala Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala

Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala64884PRTCitrobacter bacteriophage SH1 64Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Val Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ser Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Asp Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala65884PRTSalmonella bacteriophage phiSG-JL2 65Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Ala Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Val Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ala Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Val Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Leu Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Lys Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala66884PRTYersinia bacteriophage phiYeO3-12 66Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Thr Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Thr Lys 85 90 95Lys Gly Arg Lys Pro Val

Ala Tyr Ala Pro Leu Gln Ser Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ala Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Ala His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Asn Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala67884PRTEnterobacteria bacteriophage T7M 67Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Lys Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Thr Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Ser Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ala Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Leu Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Thr His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795 800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Ser 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala68884PRTEnterobacteria bacteriophage T3 68Met Asn Ile Ile Glu Asn Ile Glu Lys Asn Asp Phe Ser Glu Ile Glu1 5 10 15Leu Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Ser Ala 20 25 30Leu Ala Lys Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Leu Gly 35 40 45Glu Arg Arg Phe Leu Lys Met Leu Glu Arg Gln Ala Lys Ala Gly Glu 50 55 60Ile Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ala Thr Leu Leu Pro65 70 75 80Lys Leu Thr Thr Arg Ile Val Glu Trp Leu Glu Glu Tyr Ala Ser Lys 85 90 95Lys Gly Arg Lys Pro Ser Ala Tyr Ala Pro Leu Gln Leu Leu Lys Pro 100 105 110Glu Ala Ser Ala Phe Ile Thr Leu Lys Val Ile Leu Ala Ser Leu Thr 115 120 125Ser Thr Asn Met Thr Thr Ile Gln Ala Ala Ala Gly Met Leu Gly Lys 130 135 140Ala Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala145 150 155 160Lys His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg His Gly 165 170 175Gln Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Ile 180 185 190Gly Arg Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asp Lys Glu 195 200 205Thr Thr Met His Val Gly Ile Arg Leu Ile Glu Met Leu Ile Glu Ser 210 215 220Thr Gly Leu Val Glu Leu Gln Arg His Asn Ala Gly Asn Ala Gly Ser225 230 235 240Asp His Glu Ala Leu Gln Leu Ala Gln Glu Tyr Val Asp Val Leu Ala 245 250 255Lys Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys 260 265 270Val Val Pro Pro Lys Pro Trp Val Ala Ile Thr Gly Gly Gly Tyr Trp 275 280 285Ala Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys 290 295 300Gly Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala305 310 315 320Val Asn Leu Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu 325 330 335Ala Val Val Asn Glu Ile Val Asn Trp Lys Asn Cys Pro Val Ala Asp 340 345 350Ile Pro Ser Leu Glu Arg Gln Glu Leu Pro Pro Lys Pro Asp Asp Ile 355 360 365Asp Thr Asn Glu Ala Ala Leu Lys Glu Trp Lys Lys Ala Ala Ala Gly 370 375 380Ile Tyr Arg Leu Asp Lys Ala Arg Val Ser Arg Arg Ile Ser Leu Glu385 390 395 400Phe Met Leu Glu Gln Ala Asn Lys Phe Ala Ser Lys Lys Ala Ile Trp 405 410 415Phe Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met 420 425 430Phe Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala 435 440 445Lys Gly Lys Pro Ile Gly Glu Glu Gly Phe Tyr Trp Leu Lys Ile His 450 455 460Gly Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile465 470 475 480Ala Phe Ile Glu Lys His Val Asp Asp Ile Leu Ala Cys Ala Lys Asp 485 490 495Pro Ile Asn Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe 500 505 510Leu Ala Phe Cys Phe Glu Tyr Ala Gly Val Thr His His Gly Leu Ser 515 520 525Tyr Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile 530 535 540Gln His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val545 550 555 560Asn Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala 565 570 575Gln Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Pro 580 585 590Asn Glu Met Ile Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu 595 600 605Lys Leu Lys Leu Gly Thr Ser Thr Leu Ala Gln Gln Trp Leu Ala Tyr 610 615 620Gly Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr625 630 635 640Gly Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Asp Asp Thr Ile 645 650 655Gln Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn 660 665 670Gln Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val 675 680 685Thr Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala 690 695 700Lys Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu705 710 715 720Arg His Arg Cys Ala Val His Trp Thr Thr Pro Asp Gly Phe Pro Val 725 730 735Trp Gln Glu Tyr Arg Lys Pro Leu Gln Lys Arg Leu Asp Met Ile Phe 740 745 750Leu Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Leu Lys Asp Ser 755 760 765Gly Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val 770 775 780His Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His785 790 795

800Glu Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly 805 810 815Thr Ile Pro Ala Asp Ala Gly Lys Leu Phe Lys Ala Val Arg Glu Thr 820 825 830Met Val Ile Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Ser 835 840 845Gln Phe Ala Asp Gln Leu His Glu Thr Gln Leu Asp Lys Met Pro Pro 850 855 860Leu Pro Lys Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp865 870 875 880Phe Ala Phe Ala69883PRTArtificial SequenceSynthetic construct 69Met Asn Ile Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Ile Leu Ala Asp His Tyr Gly Ala Gln Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ala Tyr Glu Glu Gly Glu 35 40 45Lys Arg Phe Leu Lys Met Leu Glu Arg Gln Ile Lys Ala Gly Glu Phe 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Leu Ser Thr Leu Leu Pro Lys65 70 75 80Leu Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Ala Ala Lys Arg 85 90 95Gly Lys Lys Pro Val Ala Tyr Asn Pro Leu Gln His Val Lys Pro Glu 100 105 110Ala Ala Ala Phe Ile Thr Leu Lys Val Thr Leu Ala Cys Leu Thr Lys 115 120 125Ala Glu Phe Thr Thr Ile Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Thr Lys Glu Glu 195 200 205Ser Ile His Val Gly Val Arg Met Leu Glu Leu Leu Ile Glu Ser Thr 210 215 220Gly Leu Val Glu Leu His Arg Pro Asn Ala Gly Asn Val Gly Lys Asp225 230 235 240Val Glu Met Ile Gln Leu Ala Pro Glu Tyr Val Asp Leu Leu Ala Lys 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Tyr Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Thr Ser Ile Val Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Val305 310 315 320Asn Ile Ala Gln Asn Thr Pro Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Val Asn Glu Ile Val Asn Trp Lys His Cys Pro Val Ala Asp Val 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Pro Lys Pro Glu Asp Ile Asp 355 360 365Thr Asn Glu Ala Ala Leu Lys Ala Trp Lys Lys Ala Ala Ala Ala Ile 370 375 380Tyr Arg Lys Asp Lys Ala Arg Val Ser Arg Arg Leu Ser Met Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn Phe Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Asp Gly Phe Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Glu Asn Ile Met Ala Cys Ala Lys Asp Pro 485 490 495Leu Asn Asn Glu Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Arg Asp Glu Ile Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Asp 565 570 575Lys Val Asn Glu Ile Leu Lys Gln Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Glu Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Thr Glu Lys 595 600 605Leu Lys Leu Gly Thr Lys Glu Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Lys Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Val Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Arg Lys Pro Val Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val His Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Gly Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asn Thr Tyr Glu Asp Asn Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala70883PRTArtificial SequenceSynthetic construct 70Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu1 5 10 15Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45Lys Arg Phe Leu Lys Met Leu Glu Arg Gln Val Lys Ala Gly Glu Ile 50 55 60Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys65 70 75 80Leu Thr Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Ala Ala Lys Arg 85 90 95Gly Lys Arg Pro Val Ala Tyr Gln Pro Leu Gln Gly Ile Lys Pro Glu 100 105 110Ala Val Ala Phe Ile Thr Ile Lys Val Val Leu Ala Ser Leu Thr Ser 115 120 125Ala Asp Asn Thr Thr Ile Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys145 150 155 160His Phe Lys Lys His Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp Asn Lys Glu Glu 195 200 205Ser Met His Val Gly Ile Arg Met Ile Glu Met Leu Ile Glu Ser Thr 210 215 220Gly Leu Val Glu Leu His Arg His Asn Ala Gly Val Val Gly Gln Asp225 230 235 240Ser Glu Thr Ile Gln Leu Ala Pro Glu Tyr Val Glu Ala Leu Ala Lys 245 250 255Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270Val Pro Pro Lys Pro Trp Val Ser Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Val305 310 315 320Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335Val Val Asn Glu Ile Val Asn Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350Pro Ala Ile Glu Arg Glu Glu Leu Pro Pro Lys Pro Asp Asp Ile Asp 355 360 365Thr Asn Glu Glu Ala Leu Lys Ala Trp Lys Lys Ala Ala Ala Ala Val 370 375 380Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe385 390 395 400Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Pro Met Phe 420 425 430Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445Gly Lys Pro Ile Gly Lys Glu Gly Phe Tyr Trp Leu Lys Ile His Gly 450 455 460Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys465 470 475 480Phe Ile Glu Asp Asn His Asp Asn Ile Met Ala Cys Ala Lys Asp Pro 485 490 495Leu Asp Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn545 550 555 560Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Asp 565 570 575Lys Val Asn Glu Ile Leu Lys Gln Asp Val Ile Asn Gly Thr Asp Asn 580 585 590Glu Val Val Thr Val Thr Asp Lys Asp Thr Gly Glu Ile Ser Glu Lys 595 600 605Leu Lys Leu Gly Thr Lys Glu Leu Ala Gln Gln Trp Leu Ala Tyr Gly 610 615 620Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly625 630 635 640Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Asp Ala Val Ser Val Thr 675 680 685Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Lys Glu Ile Leu Arg705 710 715 720Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735Gln Glu Tyr Arg Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780Ser Gln Asp Gly Ser His Leu Arg Met Thr Val Val Tyr Ala His Glu785 790 795 800Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815Ile Pro Ala Asp Ala Gly Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830Val Asn Thr Tyr Glu Asn Asn Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860Pro Ala Lys Gly Asn Leu Asn Leu Gln Asp Ile Leu Lys Ser Asp Phe865 870 875 880Ala Phe Ala7149DNAArtificial SequenceSynthetic construct 71aattctaata cgactcacta tagggagagg cccggcatgt ggtgcataa 497223DNAArtificial SequenceSynthetic construct 72cagtatgcca agaccgactc aga 237334DNAArtificial SequenceSynthetic construct 73cgtacgagaa gaggaagccc aagagccacg tacg 347418DNAArtificial SequenceSynthetic construct 74taatacgact cactatag 187518DNAArtificial SequenceSynthetic construct 75aattaaccct cactaaag 187618DNAArtificial SequenceSynthetic construct 76tatttaccct cactaaag 187723DNAArtificial SequenceSynthetic constructmisc_feature(22)..(22)n is a, c, g, or t 77atttaggtga cactatagaa gng 23

Claims

1. An RNA polymerase, wherein the RNA polymerase: (a) has at least 90% sequence identity to SEQ ID NO:1; and (b) comprises at least one amino acid substitution corresponding to a position selected from the group consisting of: 109, 205, 534, 567, and 618 of SEQ ID NO:1.

2. An RNA polymerase of claim 1, wherein the RNA polymerase comprises at least two substitutions at positions corresponding to positions selected from the group consisting of: 109, 205, 388, 534, 567, and 618 in SEQ ID NO:1.

3. An RNA polymerase of claim 1, wherein the RNA polymerase comprises substitutions at positions corresponding to positions 109, 205, 534, 567, and 618 in SEQ ID NO:1.

4. An RNA polymerase of claim 1, wherein the substitution corresponding to position 109 of SEQ ID NO:1 is I109L, the substitution corresponding to position 205 of SEQ ID NO:1 is H205S, the substitution corresponding to position 534 of SEQ ID NO:1 is L534V, the substitution corresponding to position 567 of SEQ ID NO:1 is V567P and the substitution corresponding to position 618 of SEQ ID NO:1 is G618Q.

5. An RNA polymerase of claim 1, further comprising at least one additional amino acid substitution at position corresponding to a position selected from the group consisting of: 75, 83, 108, 206, 227, 281, 297, 312, 323, 327, 333, 340, 354, 362, 375, 388,428, 446, 454, 461, 495, 510, 584, 591, 642, 711, 724, 740, 788, 832, 834, 835, 843, 847, 849, 856, 863, 866 and 877 of SEQ ID NO:1.

6. An RNA polymerase of claim 1, comprising at least 10 additional substitutions at positions corresponding to a position selected from the group consisting of: 75, 83, 108, 206, 227, 281, 297, 312, 323, 327, 333, 340, 354, 362, 375, 788, 428, 446, 454, 461, 495, 510, 584, 591, 642, 711, 724, 740, 788, 832, 834, 835, 843, 847, 849, 856, 863, 866 and 877 of SEQ ID NO:1.

7. An RNA polymerase of claim 1, comprising at least one additional amino acid substitution selected from the group consisting of: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, D388E, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K and E877R.

8. An RNA polymerase of claim 1, comprising at least ten additional substitutions selected from the group consisting of: T75Q, A83K, E108L, K206P, V227I, I281P, V297I, Y312D, A323I, A327P, K333P, V340E, A354Q, M362P, T375K, T375N, D388E, A428P, L446F, K454P, K461R, S495N, C510Q, A584K, D591E, K642R, K711R, A724P, K740R, G788A, M832F, D834E, T835L, A843Q, D847E, F849V, S856T, A863P, A866K and E877R.

9. An RNA polymerase of claim 1, wherein, as a result of the at least one amino acid substitution, the RNA polymerase has increased activity at temperatures above 42.degree. C., 45.degree. C., 50.degree. C. or 55.degree. C. relative to the T7 RNA polymerase of SEQ ID NO:1.

10. An RNA polymerase of claim 1, wherein the polymerase has at least 95% sequence identity to any of SEQ ID NOs:52-70.

11. A fusion protein comprising: the RNA polymerase of claim 1; and a DNA binding domain.

12. A composition comprising: the RNA polymerase of claim 1; and a buffering agent.

13. A composition of claim 12, further comprising ribonucleoside triphosphates and/or a modified nucleotide.

14. A composition of claim 12, further comprising glycerol, salt, EDTA, and/or detergent.

15. A composition of claim 12, further comprising a template DNA molecule comprising: a bacteriophage RNA polymerase promoter, operably linked to a target nucleotide sequence to be transcribed.

16. A composition of claim 12, further comprising a temperature sensitive inhibitor of the RNA polymerase.